April 2002
Volume 43, Issue 4
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Eye Movements, Strabismus, Amblyopia and Neuro-ophthalmology  |   April 2002
Transcriptional Profile of Rat Extraocular Muscle by Serial Analysis of Gene Expression
Author Affiliations
  • Georgiana Cheng
    From the Departments of Ophthalmology,
  • John D. Porter
    From the Departments of Ophthalmology,
    Neurology, and
    Neurosciences, Case Western Reserve University; and The Research Institute of University Hospitals of Cleveland, Cleveland, Ohio.
Investigative Ophthalmology & Visual Science April 2002, Vol.43, 1048-1058. doi:
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      Georgiana Cheng, John D. Porter; Transcriptional Profile of Rat Extraocular Muscle by Serial Analysis of Gene Expression. Invest. Ophthalmol. Vis. Sci. 2002;43(4):1048-1058.

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      © 2016 Association for Research in Vision and Ophthalmology.

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Abstract

purpose. Although extraocular muscle (EOM) is skeletal muscle, aspects of its biology are unlike other striated muscle. In this study, the broad molecular genetics profile underlying the novel EOM phenotype was examined.

methods. Serial analysis of gene expression (SAGE) was used to quantify adult rat EOM gene transcripts. SAGE isolates and sequences 10-bp tags from defined locations in mRNA-derived cDNA. Tag sequence-location was used to extract transcript identity from a curated SAGE database, and detection frequencies reflected abundance of corresponding mRNAs.

results. The 54,764 expressed sequence tags generated and sequenced from EOM included 17,602 unique tags. Of the unique tags, 7.8% were detected at high to intermediate levels (≥5 copies), 19.3% at lower levels (2–4 copies), and 72.9% as single copies; 40% of the tags matched known expressed sequence tags (ESTs), most of which (85.7%) represented a unique EST. Tags without matches in the SAGE database and those expressed as single copies only were not considered further. SAGE tags expressed at more than 0.1% of total transcripts reflected several aspects of muscle biology, including sarcomeric structure, energy metabolism, and ribosomal protein expression. Genes highly expressed in EOM were compared with other existing muscle expression databases to identify conserved and novel patterns in EOM.

conclusions. The data provide a normative gene expression database and a novel molecular signature that will facilitate study of EOM development and function and of the mechanisms behind its preferential targeting or sparing in neuromuscular disease.

The oculomotor system exhibits precision and diversity unlike that of any other skeletomotor system. As the effector organ for ocular motility, the extraocular muscles (EOMs) are adapted for novel functional demands. Thus, there are fundamental phenotypic differences between EOMs and other skeletal muscles. 1 2 3 In skeletal muscle biology, the basic structural and functional unit is the multinucleated myofiber. Patterned covariations in myofiber traits that determine contraction speed and energy metabolism (aerobic, anaerobic, or both) identify distinctive muscle fiber types. Consensus classification schemes have three to four distinct fiber types, each differing in speed and fatigue resistance, that are conserved in virtually all mammalian skeletal muscles. 4 5 Functional role-specific skeletal muscles thus contain the requisite proportions of these few myofiber types. EOM is a clear exception to this rule, because the six EOM-specific fiber types do not fit any of the traditional skeletal muscle fiber classification schemes. 2 3  
Gene-profiling technologies have demonstrated considerable power in generation of cell and tissue molecular signatures and identification of disease-associated gene expression changes and represent a rapid and efficient means to elucidate alterations in cell signaling or metabolic pathways. Despite this potential, there has been only limited use of genome-wide screening techniques in vision science. We recently gained new insight into the underlying molecular mechanisms that differentiate EOMs from other skeletal muscles through pair-wise muscle group comparisons, using high-density oligonucleotide microarray technology. 6 These data established striking similarities in gene expression patterns between mouse hindlimb and some craniofacial (masticatory) muscles, but substantial divergence of EOMs from both muscle groups. Although the scope of our prior DNA microarray study was broad—approximately 10,000 genes and ESTs evaluated—this sample represents, at best, no more than 25% to 35% of the mouse genome. Thus, there indeed are other as yet unrecognized patterned differences in gene expression between EOMs and prototypical skeletal muscles. Collectively, knowledge of the molecular signature of EOMs is vital for both modeling of its novel structural and functional properties and for understanding its differential response in a variety of neuromuscular and autoimmune diseases. 3 7 8 9 10 11 12 13 14  
For this study, we used serial analysis of gene expression (SAGE) to take advantage of recently available genome project databases and high-throughput DNA sequencing technology to obtain a quantitative transcriptional profile of EOM. In brief, SAGE relies on the isolation of a short oligonucleotide sequence from a defined location within cDNA derived from tissue poly(A)+ RNA to quantify the specific transcript. 15 The restriction enzyme, NlaIII, termed the “anchoring” enzyme in SAGE, cuts at the 3′-most occurrence of an NlaIII restriction site. After product ligation to a linker, a tagging enzyme is used to release a 10-bp SAGE tag. The location and sequence of the tag identifies a specific transcription product. Often SAGE tags map to only one expressed gene, but a caveat to the technique is that tag-to-gene assignments are sometimes ambiguous and a single tag may map to two or more genes. Accumulation of thousands of SAGE tags specific for genes or ESTs (termed matched tags) allows the establishment of a global gene expression signature for a tissue. SAGE is differentiated from the other contemporary gene-profiling technology, DNA microarray, in that it is not restricted to a preselected set of probes, but has the potential to detect any expressed transcript. Any 10-bp SAGE tag sequences that do not match to known genes or ESTs (novel tags), but are present at high copy numbers, represent logical targets for gene discovery. 
Using a modification of SAGE adapted for small amounts of starting mRNA (microSAGE 16 ) we profiled more than 50,000 transcripts from adult rat EOM, to generate a tissue-specific expression library, and compared these data with previously published skeletal muscle profiles. The data established that EOM has a unique molecular signature. 
Materials and Methods
Animals
These studies used 45-day-old Sprague-Dawley rats (Harlan, Indianapolis, IN). All animal procedures were approved by the Institutional Animal Care and Use Committee at Case Western Reserve University and were in accordance with the ARVO Statement for the Use of Animals in Ophthalmic and Vision Research. EOMs were rapidly isolated and removed from five male rats after asphyxiation with carbon dioxide. Tissues were flash-frozen in liquid nitrogen and stored at −80°C. 
SAGE Protocol
In this study, we used the published microSAGE protocol (version 1.0e; available at http://www.sagenet.org/), 16 with modifications that will be described later. EOMs from five rats were combined, 50 mg of tissue was homogenized, and mRNA was isolated using an extraction kit (Dynabeads mRNA Direct; Dynal, Oslo, Norway), according to the manufacturer’s instructions. Poly(A)+ mRNA was bound to oligo (dT) 25 and converted to cDNA with a synthesis kit (Gibco BRL, Rockville, MD), according to the manufacturer’s instructions. The cDNA was evaluated by PCR for integrity and then cleaved with the anchoring enzyme, NlaIII (New England BioLabs, Beverly, MA). The identity of original mRNA copies is derived from the sequence of SAGE tags lying immediately 3′ to the 3′-most NlaIII cDNA-cleavage site. The 3′ terminal cDNA fragments were isolated with magnetic beads and ligated to one of two annealed linker pairs. Ligated linker contained recognition sites for BsmFI, allowing cDNA- tags to be released from beads by the tagging enzyme BsmFI (New England BioLabs). Tags then were ligated to one another to form ditags, after blunt ending with Klenow (Pharmacia, Peapack, NJ). 
The ditags were amplified by PCR and products analyzed by polyacrylamide gel electrophoresis (PAGE) and then digested with NlaIII. The products containing the ditags were then ligated to form concatamers. To prevent overrepresentation of clones with short insertions (which ligate to the vector more efficiently than larger ones), the concatamers were size fractionated by PAGE. Three separate fractions, approximately corresponding to 0.5 to 0.9 kb, 0.9 to 1.5 kb, and 1.5 to 2.5 kb, were cut from gels. The concatamer-containing cDNA from each fraction was purified and cloned into the SphI site of a vector (pZErO-1; Invitrogen, Carlsbad, CA). Insert-containing colonies were screened by PCR with M13 forward and reverse primers. PCR products containing inserts of more than 616 bp in length, which should contain at least 15 ditags each, were submitted for DNA sequencing. 
SAGE Tag Sequencing and Analysis
SAGE tag sequencing was performed by San Ming Wang (Department of Hematology and Oncology, University of Chicago Medical Center, Chicago, IL) with a sequencing kit and sequencer (Big-Dye Sequencing Kit and Prism 377 DNA Sequencer; (PE-Applied Biosystems, Foster City, CA). Tag sequences were extracted from clone sequence data with SAGE 300 software. Conversion of tag sequence data to represented genes was performed using software at the Rn.seq.all.z/SAGE map site, available at the National Center for Biotechnology Information (NCBI; http://www.ncbi.nlm.nih.gov). All sequences were arranged in a FASTA file, and the NCBI Unigene database (available at http://www.ncbi.nlm.nih.gov/Unigene) was searched to yield identities of the original mRNA. This tag-to-gene screen separates the 10-bp tags with matches to known genes and ESTs (matched tags) from those with no matches (novel tags) and quantifies tag frequency. 
EOM SAGE data were compared with human leg muscle gene expression frequency data in existing SAGE (http://www.urmc.rochester.edu/smd/CRC/Swindex.html) and EST (http://telethon.bio.unipd.it/GETProfiles/) databases. The statistical test of Audic and Claverie, 17 available as an executable program via a Web interface (http://igs-server.cnrs-mrs.fr/∼audic/significance.html), was used to identify genes that were differentially expressed in EOM versus leg muscle. This digital gene expression profiling method compares databases by computing probabilities based on individual tag copy numbers and overall SAGE library size. 
Results
Distribution of SAGE Tags
SAGE was used to generate a quantitative gene expression profile from EOM. Two quality control measurements indicate that the SAGE data presented herein are of high quality. First, linker contamination of our library was very low; our data contained only 0.086% linker sequences versus 0.65% or more in other recent studies. 18 19 Second, analysis of the average GC content 20 of tags in our SAGE library (48.04% GC) indicated no GC content bias (bias in a library can arise from denaturation and experimental loss of AT-rich regions and is defined as GC content ≥55%). Because quantitative differential gene expression methodologies are highly sensitive to the quality of both the original mRNA preparation and the cDNA library, quality control measures are an important consideration. 
SAGE generated two data sets from rat EOM: 10-bp tags for qualitative identification of expressed transcripts when matched against the NCBI SAGE database and relative transcript abundance data that provide for both a global expression profile and the ability to make quantitative same-gene and cross-gene expression frequency comparisons. A total of 17,602 unique SAGE tag sequences were extracted from the 54,764 expressed sequences we accumulated from adult EOM. The finding of approximately 32% unique tags in EOM was higher than reported in a prior skeletal muscle SAGE study (∼23%). 21 Individual EOM-derived tag abundance ranged from 1 to 2109 copies per gene. Among the 17,602 identified unique tags, 72.9% were present as single copies; 19.3% had 2 to 4 copies; 4.3%, 5 to 9; 3.2%, 10 to 99; and 0.3% 100 or more (Fig. 1) . Tags expressed only as single copies were not considered further. 
Most Highly Expressed Genes in EOM
The frequency of occurrence of each SAGE tag reflects the relative abundance of the corresponding mRNA. Both intergene and same-gene abundance comparisons are valid with SAGE data, because tags are generated without any selection bias, and the in vivo frequency of an mRNA directly determines the probability of a tag’s appearing in a SAGE library. Comparison of the 17,602 unique tags seen in EOM to the NCBI SAGE database showed that 40.4% of the tags matched known expressed sequences, including both known genes and ESTs, and 59.6% of all tags had no match. Analysis of matched and novel tags showed that tags present in high copy numbers also had a high percentage of matches to known expressed sequences, whereas the majority of the novel tags were concentrated in the low-abundance class and were especially frequent among the single-copy group (Fig. 1) . The novel (unmatched) tags were not considered further. 
Table 1 shows the identities of the most abundant SAGE tag species in rat EOM (n = 93), representing all genes and ESTs expressed at a level 0.1% or more of total tags accumulated (i.e., ≥55 copies). This group is identified throughout as the most frequently expressed genes in EOM. All but 8 of the 93 tags in this group were matched tags, with 10-bp sequences identical with at least one tag in the NCBI SAGE database (Fig. 1) . Comparison of our SAGE data with an existing DNA microarray database from rat EOM (Khanna S, Merriam A, Leahy P, Andrade F, Porter J, unpublished data, 2001) showed that many transcripts detected by SAGE have not been previously identified in EOM. Of the 93 most frequently expressed transcripts, DNA microchip data were available for 55 of them; 48 of the SAGE transcripts had been previously identified as present, and 7 were judged as absent in rat EOM by the microarrays (∼13% noncorrespondence; Table 1 ; see the Discussion section). The two most abundant mRNA species in EOM code for the Glu-Pro dipeptide repeat protein (present as four different SAGE tags accounting for 3.9%, 1.0%, 0.2%, and 0.2% of total transcripts) and N G,N G-dimethylarginine dimethylaminohydrolase (Ddah1; three tags: 3.3%, 2.8%, and 0.7% of transcripts). The presence of multiple SAGE tag species for these two genes may represent alternatively spliced isoforms. 
Global Gene Expression Profile of EOM
Functional classes were assigned to all genes identified by SAGE tags with 10 or more copies. Gene classifications were organized into areas of highest relevance for muscle biology. Among the most abundant transcript classes in EOM were those associated with sarcomeric structure, energy metabolism, and ribosomal structural protein expression. Data for selected genes in some important functional categories are presented in Table 2 ; the entire EOM SAGE data set has been deposited at the NCBI public repository (http://www.ncbi.nlm.nih.gov/geo/) and is accessible under accession number GSM581. 
Differential Gene Expression in EOM and Skeletal Muscle
A prior DNA microarray study detected broad gene expression differences between mouse EOM and leg musculature. 6 Quantitative expression comparisons are the best means of gaining insight into mechanisms that underlie phenotypically different muscle groups. However, there are only a few skeletal-muscle–profiling studies in which SAGE has been used, 21 22 23 24 limiting the current availability of data for evaluation of muscle group gene expression differences. All but one of the prior skeletal muscle SAGE studies has been in humans, and the one rat skeletal muscle SAGE library 24 may be too small (∼2256 total leg muscle tags, whereas 50,000 is generally regarded as optimal), because sample size interacts with expression frequency in statistically stringent muscle group comparisons. Thus, we compared gene expression levels between the most frequently expressed genes in ratEOM to human skeletal muscle data in an existing SAGE library (vastus lateralis muscle) (http://www.urmc.rochester.edu/smd/CRC/Swindex.html) 21 (Tables 2 3) 3 . Of the most frequently expressed genes in EOM, only 43% were also represented in the human SAGE skeletal muscle library, often at lower levels, and of the top 10 EOM transcripts only myosin light chains 1 and 2 and myoglobin were represented in the skeletal muscle SAGE library at any reported level. In the converse comparison, of the top 10 transcripts in the skeletal muscle SAGE database, 5 were highly expressed in EOM (α-actin, glyceraldehyde 3-phosphate dehydrogenase, fructose bisphosphate aldolase A, ubiquinone oxidoreductase chain 3 [URF3], and muscle creatine kinase), whereas 5 were not (cytochrome c oxidase 2, β-cardiac myosin, NADH-and ubiquinone oxidoreductase chain 4 [URF4], β-tropomyosin, and proteosome 26S subunit [ATPase 6]). 
Databases with information about the abundance of identified ESTs in skeletal muscle libraries represent an additional source of quantitative muscle gene transcription data. Use of such EST databases for muscle group comparisons relies on the assumption that the frequency of gene-specific ESTs in non-normalized, nonsubtracted libraries is an indicator of the transcriptional activity of that gene. We compared the most frequently expressed genes (≥0.1% of transcripts; n = 177 genes with ≥30 ESTs/gene) in a published adult human skeletal muscle EST database (http://telethon.bio.unipd.it/GETProfiles/) 25 to our group of the most frequently expressed genes in rat EOM (n = 93 genes). The modest degree of correspondence in muscle group transcription patterns was apparent in the finding that only 32% of the most frequently expressed genes in rat EOM also were highly expressed in the human vastus lateralis EST database (Tables 2 3) 3 . Of the five most expressed genes in the human EST database, three were also among the most frequently expressed genes in rat EOM (muscle creatine kinase, myoglobin, and glyceraldehyde-3-phosphate dehydrogenase) and two were not (ribosomal protein L37a and slow troponin T1). 
Discussion
Skeletal muscle is an elegant structure–function model. Individual myofibers exhibit patterned variations in traits that collectively determine whether contractions are slow or fast and whether the most efficient mode of energy metabolism is matched to the fiber usage pattern. Although factors directing their development are only poorly understood, 26 27 28 29 the fundamentally distinct properties of EOM are likely a consequence of the diverse demands placed on them by oculomotor control systems with dynamic ranges from slow vergence movements to rapid saccades. Although there is not an absolute one-to-one correspondence of transcript level to protein content, the biological potential of EOM, like any tissue, is determined at the gene-expression level. In this study, we used a powerful gene profiling tool, SAGE, to define a molecular signature for EOM that is unbiased by any predetermined gene selection criteria. Strict quality control measures indicate low to absent contamination or selective loss of transcripts, thereby providing confidence in these results. Data establish the molecular expression profile of EOM and, when compared with the expression pattern of other skeletal muscles, strongly support the notion that EOM is a fundamentally different type of skeletal muscle. 6  
SAGE as a Gene Expression–Profiling Tool
There are two contemporary tools for determination of broad gene expression patterns, DNA microarray and SAGE, each with its own particular advantages. Combined use of both gene-profiling technologies has allowed important advances in understanding retinal cell lineage. DNA microarray relies on either cDNA or oligonucleotide probes arrayed on a microchip and the differential binding of labeled target tissue transcripts to the probe arrays. Microarray analysis has high sensitivity and is equally efficient at detection of both high and low copy number transcripts, but its use is restricted to only those gene-specific probes that have been designed into an array. Although high-density commercial arrays can evaluate expression changes using more than 10,000 probes, this is still a restricted sample of the entire mammalian genome, and data obtained from low-density or specialized arrays are inherently biased by probe quality and selection. DNA microarray is excellent, however, at quantification of same-gene changes in expression level but does not provide a portable measure of the level of gene expression and does not allow reliable cross-gene comparisons due to differences in transcript-to-probe binding efficiency. 
By contrast, SAGE can extract and identify the transcript of any nuclear-transcribed gene, without preexisting bias by investigator or microarray chip designer and allows equally precise quantification of both same-gene and cross-gene expression levels. Moreover, SAGE data are portable in that a database is universally acceptable for comparisons in other experiments, whereas microarray data sets may be too dependent on factors such as microarray type, probe design, transcript isolation, purification, labeling and hybridization conditions, and scanner photomultiplier tube sensitivity to be useful in direct quantitative comparisons with data obtained by other investigators. However, low-copy-number mRNAs are poorly detected by SAGE, in part because high-copy-number transcripts overwhelm rare transcripts during SAGE tag isolation. Signal transduction and transcription factor transcripts, in particular, can be specifically detected in microarray experiments, but are generally expressed at levels too low to be consistently found in a library of 50,000 SAGE tags. In sum, SAGE quantifies the more highly expressed gene transcripts, allowing them to be placed in a relative rank order and compared with other SAGE databases, without the bias of investigator or commercially selected probes. 
EOM and Tissue Expression Analysis
EOM exhibits a complex architecture, containing six diverse muscle fiber types plus cellular elements associated with an extensive extracellular matrix and vascular supply (for review, see Spencer and Porter 2 ). As potential transcript sources, the predominant cell types in skeletal muscle include myofibers, muscle stem (satellite) cells, fibroblasts, Schwann cells, resident cellular defense elements, endothelial cells, circulating cells trapped in the microvasculature, and arteriole-associated smooth muscle cells. Yet, myonuclei comprise an estimated 75% of all nuclei in skeletal muscle tissue. 30 Thus, it is a reasonable assumption that our SAGE data predominantly represent EOM transcripts. The substantial level of myofibrillar proteins exclusively expressed in muscle and electron transport proteins heavily concentrated in muscle is consistent with muscle transcripts as the predominant species of mRNA in the EOM. An exception is the presence of high copy numbers of β- and α-globin (collectively, 0.8% of total SAGE tags in EOM), suggesting that blood elements contribute to the transcript pool. 
EOM myofibers are specialized for low force production but high fatigue resistance. Consistent with observations that myofilaments occupy less of the EOM fiber cross-sectional area than in other skeletal muscles, 31 myofibrillar protein transcripts (e.g., myosin, α-actin, troponin, and tropomyosin) comprised only 10.6% of EOM gene transcripts detected at copy numbers higher than 100, versus the 27.8% myofibrillar transcripts in the SAGE human leg muscle library. 21 Likewise, EOM has among the highest mitochondrial content of any skeletal muscle (among the six fiber types, 5%–24% of fiber volume). 31 The abundance of mitochondria is reflected in the relative abundance of nuclear-encoded transcripts for oxidative phosphorylation proteins in EOM (mitochondrial transcripts are not isolated by the methodology used herein and were not detected in our library). Taken together, the global gene expression profile obtained by SAGE, to a great extent, specifies the transcriptional properties of the muscle fiber component of EOM and faithfully reflects myofiber subcellular organelle content. 
The EOM Transcriptome Versus That of Other Skeletal Muscles
Although the rat genome project is not complete (and thus the NlaIII restriction sites necessary for SAGE tag identification have not been sequenced for all genes), very few (8.6%) of the most frequently expressed genes in EOM did not have a corresponding tag identifier in the NCBI rat SAGE database (Table 1) . Thus, the highly expressed genes detected and quantified by SAGE can be confidently compared with other existing skeletal muscle data without the limitation of excessive numbers of unknown transcripts. Of the top 0.1% or more of EOM transcripts, we had previously detected 52% of them as present in rat EOM by DNA microarray (Khanna S, Merriam A, Leahy P, Andrade F, Porter J, unpublished data, 2001). The remainder of the genes either were not represented on the microarray chips or the array probes do not appear to adequately detect the intended genes (e.g., muscle creatine kinase and fructose bisphosphate aldolase are established muscle genes but were not detected by the specific probe sets on Affymetrix rat microarrays obtained from Affymetrix (Santa Clara, CA); Khanna S, Merriam A, Leahy P, Andrade F, Porter J, unpublished data, 2001). Our comparisons of the EOM transcription profile with that of the human vastus lateralis muscle, a prototypical skeletal muscle often used in diagnostic muscle biopsies, showed that their concordance in expression pattern was only 33% to 47% (Table 3) . 21 25 A caveat of this analysis is the possibility of species differences that may distort observed muscle class differences. Normally, functionally distinct skeletal muscles differ only in their percentage content of the three to four highly conserved myofiber types. This translates into relatively small gene expression differences between predominantly fast- and slow-twitch skeletal muscles. 32 In that study, detected differences were largely restricted to genes that could be easily associated with the known fiber type differences. By contrast, EOM fiber types are unlike the traditional skeletal muscle types, and the expression differences between EOM and other skeletal muscles seen here extend well beyond genes that specify fast- or slow-twitch fiber phenotypes. 
The two most frequent transcripts in EOM, Glu-Pro dipeptide repeat protein and Ddah1, were not among the most frequent transcripts in either of the existing vastus lateralis muscle databases. Because the role of the dipeptide repeat proteins is unknown, it is difficult to speculate about the functional significance of this difference in gene expression. Proteins with glutamic acid-proline are particularly abundant in cardiac and skeletal muscle 33 and have been previously detected as present in EOM (Table 1 and unpublished data, 2001). Ddah1 codes for an enzyme that metabolizes neuronal nitric oxide synthase (nNOS) inhibitors. High Ddah1 levels in EOMs may provide for selective regulation of NOS inhibitor concentration and, in turn, positive regulation of NOS activity in vitro. 34 The muscle group–specific role that specialized control of nNOS might play in EOM is unknown, but the high expression level of Ddah1 suggests that it is vital to EOM function, perhaps by modulating EOM contractile function in ways suggested by Richmonds and Kaminski. 35  
Some of the differences between our EOM SAGE data and the human skeletal muscle EST library likely are related to known functional differences between the two muscles. Expression mismatches can be attributed, in part, to the fiber type composition of the two muscles (type IIA and I predominance in the vastus lateralis versus novel fiber types, with no type I [slow-twitch] component, in EOM). Although the two muscle groups shared high expression of at least five key sarcomeric genes (α-1 actin; myosin heavy chain 1 [type IIX/D]); troponin I, fast; tropomyosin 1; and myosin light chain 1), seven of the most frequently expressed skeletal muscle sarcomeric proteins not detected among the most highly expressed group in EOM were slow-twitch muscle fiber–specific transcripts (β-cardiac [slow] myosin heavy chain; troponin I, slow; myosin light chain 3, slow; myosin-binding protein C, slow-type; myosin light chain, regulatory, slow; troponin C, slow; sarcoplasmic reticulum calcium ATPase 2 [slow-twitch]). The fast-twitch type IIA muscle fiber transcript (myosin heavy chain 2) does not have a rat SAGE tag identifier and thus its levels could not be evaluated. The relative low number of slow-twitch transcripts relates to the total absence of slow-twitch fiber types in EOM; some slow-twitch transcripts may be represented in the novel slow-tonic EOM fiber types that comprise a small percentage of EOM fibers. By contrast, the EOM-specific myosin heavy chain (Myh13; 0.2% of transcripts) is not expressed in limb musculature. By SAGE, myosin heavy chain expression levels in EOM were in the order EOM > IIX > IIB > embryonic, with three other isoforms known to be present in EOM (neonatal, I, and IIA) not detected at more than 10 copies. A quantitative PCR study ranked the frequency of myosin transcripts in rat EOM in the order IIB > IIA > IIX > EOM > I > neonatal > embryonic. 36 Some, but not all, interexperimental differences can be attributed to the lack of SAGE tag identifiers for, and therefore an inability to assess, the rat neonatal and IIA myosin heavy chain isoforms. 
Skeletal muscle fiber types differ in mechanisms for energy metabolism, with the predominant modes being oxidative, oxidative-glycolytic, and glycolytic. Most skeletal muscles are dependent on substantial glycogen stores for energy metabolism. By contrast, EOM exhibits exceptionally low glycogen content for a skeletal muscle. 2 Our prior gene profiling study showed that expression of many enzymes related to glycogen metabolism and gluconeogenesis were at lower levels than in leg musculature. 6 Our observation that glycogen phosphorylase (<0.02% of EOM transcripts) and phosphoglycerate mutase (not detected in EOM at >0.009% of transcripts) are not very abundant in EOM (Table 2) is consistent with these data. Mitochondrial content represents a distinguishing feature of several EOM fiber types, with few other skeletal muscles exhibiting such a high mitochondrial volume. 31 Consistent with a substantial reliance on oxidative phosphorylation, EOM exhibited high expression levels of several nuclear-encoded isoforms of cytochrome c when compared with the vastus lateralis muscle (Table 2) . Of nine cytochrome c oxidase isoforms detected at five or more copies in EOM, four were expressed at higher levels, three at lower levels, one was not different, and one was not detected in the vastus lateralis. Also related to energy metabolism in fast-twitch muscle fiber types, enzymes concerned with phosphocreatine metabolism (muscle creatine kinase and sarcomeric creatine kinase) did not follow the established fast-twitch pattern, but were differentially expressed in EOM compared with the vastus lateralis (Table 2) . Myoglobin expression in EOM was approximately half that of the vastus lateralis. Collectively, data suggest that EOM may use novel substrates and pathways for energy metabolism that differ from those that typify other skeletal muscles. 
Another major difference between EOM and human vastus lateralis transcriptomes was in the expression of ribosomal proteins, with 11 conserved between the most highly expressed genes in the two muscle groups, 3 specific to EOM, and 58 specific to leg muscle. Although the core of the ribosome translational unit is RNA, the ribosomal proteins are integral components, decorating the rRNA subunit cores and contributing to transfer (t)RNA binding sites, among other roles. It is difficult to speculate about the significance of these substantial differences in ribosomal protein expression, because the extent to which these muscle group differences may reflect simple species differences or may be determinants of tissue-specific translation is unclear. Moreover, some ribosomal proteins have secondary functions independent of their involvement in protein biosynthesis. Until such roles are clarified, it is difficult to interpret the substantial muscle group differences in expression of this protein group. 
Finally, we detected significant differences between EOM and other skeletal muscles in the expression of several cytoskeletal structural proteins (desmin, titin, vimentin, and nebulin; Table 2 ). Desmin, as the main intermediate filament protein in skeletal muscle, is of great importance as a part of the cytoskeleton that supports transfer of generated force to the muscle tendon. 37 That levels of desmin and other noncontractile cytoskeletal proteins are either low or possibly absent in EOM strongly suggests that cytoskeletal organization in EOM may be very different from other skeletal muscles. Such internal cytoskeletal differences may directly relate to the differential sensitivity of EOM in a variety of neuromuscular disorders, including muscular dystrophy and nemaline myopathies. 
Conclusions
SAGE has been used previously to study skeletal muscle only in very limited situations (normal human profile, aging changes, and a disuse model) but has the potential to identify the molecular determinants of tissue phenotype and disease-associated changes in those determinants. Our data provide a molecular identity for an important but little understood skeletal muscle, EOM, and support the concept that EOM is dramatically different from other skeletal muscles. A broad understanding of the biology of the final common pathway for eye movements, achievable only through gene profiling, will aid global understanding of the functions and diversity of oculomotor control mechanisms. 
 
Figure 1.
 
Frequency distribution of EOM-derived SAGE tags. Matched tags denote SAGE tag sequences that map into the NCBI database, and novel tags are those that did not have a sequence match in the database. (A) Semilog plot showing the frequency distribution of SAGE tags detected at specific abundance levels. Novel tags were most frequent among tags expressed only as single copies, and their level declined relative to matched-tag frequency, with increasing copy number. (B) Plot illustrating the frequency of matched SAGE tags. When expressed as a percentage of total tags in each bin, matched-tag frequency increased as a function of copy number.
Figure 1.
 
Frequency distribution of EOM-derived SAGE tags. Matched tags denote SAGE tag sequences that map into the NCBI database, and novel tags are those that did not have a sequence match in the database. (A) Semilog plot showing the frequency distribution of SAGE tags detected at specific abundance levels. Novel tags were most frequent among tags expressed only as single copies, and their level declined relative to matched-tag frequency, with increasing copy number. (B) Plot illustrating the frequency of matched SAGE tags. When expressed as a percentage of total tags in each bin, matched-tag frequency increased as a function of copy number.
Table 1.
 
Most Frequently Expressed SAGE Tags in EOM
Table 1.
 
Most Frequently Expressed SAGE Tags in EOM
Tag Copy No. Unigene Cluster Gene Name, *
GATGCCCCCC 2109 Rn.3979 Glu-Pro dipeptide repeat protein
ATAACACATA 1792 Rn.4241 NG,NG dimethylarginine dimethylaminohydrolase (Ddah1)
ATACTGACAC 1536 Rn.4241 NG,NG dimethylarginine dimethylaminohydrolase (Ddah1)
AATCGGAGGC 584 Rn.25727 CDK110
AGCCATCCCT 530 Rn.3979 Glu-Pro dipeptide repeat protein
TACCACCTTT 464 Rn.40120 ESTs, highly similar to rat myosin light chain 1, skeletal muscle
TAGGTACAGG 387 Rn.4241 NG,NG dimethylarginine dimethylaminohydrolase (Ddah1)
TTTGGGGAGA 364 Rn.37176 Myosin light chain 2, heart
Rn.6534 Myosin light chain 2, alkali, ventricular, skeletal, slow
Rn.54542 Myosin light chain 2
TGGGTTGTCT 361 Rn.36610 Lens epithelial protein
CTCAGGTCTC 322 Rn.40511 Myoglobin
ATGACTATTA 278 None Unknown
GAGGGTCGGA 266 Rn.9924 Troponin I, skeletal, fast 2
TGGATCCTGA 264 Rn.36966 Major beta-globin
Rn.54518 Zero beta-1 globin, exons 1 and 2
TAGGCCACAC 254 Rn.2618 Mitochondrial ATP synthase beta subunit
AGCGATTCAA 236 Rn.43378 ESTs, moderately similar to mouse URF3
GCCTTCCTCA 231 None Unknown
GATAAAACCA 227 Rn.9056 ESTs
Rn.4092 Mitochondrial adenine nucleotide translocator
TGGGCACCTG 220 Rn.3454 F1-ATPase epsilon subunit
CCCAGGCACT 212 Rn.22504 Troponin T, skeletal, fast
GCATATTGAA 211 Rn.2026 ESTs, Moderately similar to mouse cytochrome c oxidase VIIb precursor
TGACAGACGA 203 Rn.43529 ESTs, weakly similar to human calmodulin
GCCTCCAAGG 202 Rn.1925 Glyceraldehyde-3-phosphate dehydrogenase, brain
Rn.54911 Glyceraldehyde-3-phosphate dehydrogenase
Rn.39164 ESTs
CACCCATACC 192 None Unknown
CACGCCTCTC 191 Rn.55177 Alpha-globin
Rn.36966 Major beta-globin
Rn.11229 Hemoglobin, alpha 1
AAAAATCATC 190 Rn.7381 ESTs, moderately similar to mouse URF5
CTAGTCTTTG 186 Rn.34942 Ribosomal protein S29
AGGAGCTCGG 177 Rn.10325 Cytochrome c oxidase subunit VIII-H
GAAATATGTG 168 Rn.2180 ESTs, moderately similar to ATP synthase lipid-binding protein P2 precursor
TTGTTAGTGC 167 Rn.1504 Malate dehydrogenase, cytosolic
GTGGCTCACA 161 Rn.51650 ESTs
Rn.40562
Rn.53514
Rn.51174
Rn.44604
Rn.43236
Rn.52705
Rn.40249
Rn.42743
Rn.43716
TAAAATTGTA 152 Rn.32408 EST, moderately similar to human KIAA0321
Rn.3905 ESTs, highly similar to creatine kinase, sarcomeric, mitochondrial, precursor
GGCTATGTAA 136 Rn.51390 ESTs, weakly similar to mouse S43118 finger protein
Rn.3819 Decorin
Rn.20597 ESTs
TGTAATGAGA 128 Rn.52850 EST
Rn.2462 ESTs, moderately similar to mouse F1F0-ATP synthase g subunit
CGAACTCTCA 124 Rn.31991 Secreted acidic cysteine-rich glycoprotein (osteonectin)
AAATAAAACT 123 Rn.5875 Tuberous sclerosis 2
Rn.40213 ESTs
Rn.6390 Cis-Golgi p28
Rn.27383
AGCCGTCCCT 120 Rn.3979 Glu-Pro dipeptide repeat protein
AAAAAAAAAA 116 Rn.20806 ESTs
Rn.20483
Rn.45252
Rn.10764
Rn.11656
Rn.43473
Rn.11739
Rn.14962
Rn.14882
Rn.2122
(continues)
Table 1A.
 
(continued) Most Frequently Expressed SAGE Tags in EOM
Table 1A.
 
(continued) Most Frequently Expressed SAGE Tags in EOM
Tag Copy No. Unigene Cluster Gene Name, *
ATCCCTGCGC 116 Rn.10756 Creatine kinase, muscle
CCAGTCCTGG 116 Rn.3357 ATP synthase, H+ transporting, mitochondrial F0 complex, subunit c (subunit 9), isoform 1
TAACTTGGAC 115 Rn.11077 Mitochondrial cytochrome c oxidase subunit Va
TCAGGCTGCC 114 Rn.1960 Ferritin subunit H
Rn.54798 Ferritin
Rn.42678 EST, moderately similar to ferritin heavy chain
CTCCTGGACA 111 Rn.2883 ESTs, weakly similar to AF099929_1 pervin
GATCAGTCAT 108 Rn.3384 Ribosomal protein L19
Rn.40331 ESTs
GAATGACCTG 108 Rn.34334 Ribosomal protein L13a
Rn.6005 ESTs, highly similar to human 40s ribosomal protein S28
AAGATCAAGA 107 Rn.39438 Actin, alpha 1, skeletal muscle
Rn.50809 EST
Rn.958 Actin, gamma 2, smooth muscle, enteric
Rn.69
TTGGGCCAGA 104 Rn.2130 ESTs, Highly similar to mouse cytochrome c oxidase VIIc precursor
Rn.21453 ESTs
ATGCAACTAC 101 Rn.6009 ESTs, highly similar to mouse cytochrome c oxidase VIb
CAAACATCCA 99 Rn.3979 Glu-Pro dipeptide repeat protein
GGAGGGGAAG 98 Rn.43378 ESTs, moderately similar to mouse URF3
Rn.22772 ESTs
TGTTCCTCTT 97 Rn.3247 Procollagen type III, alpha 1
CGGGATCTGC 97 Rn.1817 Oligomycin sensitivity conferring protein
GGGCAACCAG 97 Rn.3472 ESTs, weakly similar to human ubiquinol-cytochrome c reductase complex ubiquinone-binding protein QP-C
GGATTTGGCC 96 Rn.55312 ESTs, highly similar to acidic ribosomal protein P2
Rn.976 ESTs, highly similar to 60S acidic ribosomal protein P2
GGATTCGGTC 94 Rn.1079 Acidic ribosomal protein P0
TAATACTCAA 94 Rn.7661 ESTs
Rn.4281 DAPIT protein
GGCCACCTCC 93 Rn.34319 Myosin, heavy polypeptide 13, skeletal muscle
CCCCATCTCA 92 Rn.2005 Parvalbumin
GAGGCTGTGG 91 Rn.12939 ESTs, weakly similar to Ste20-like kinase
Rn.9738 Phosphoglycerate mutase
TCCAATAAAG 87 Rn.973 ESTs, highly similar to 60S acidic ribosomal protein
Rn.47395 EST
AGCGCCCAGA 83 Rn.5119 Cytochrome c oxidase subunit VIa polypeptide 2, heart
TTAAGACTAG 78 Rn.25727 CDK110
TTAATAAATG 78 Rn.2528 Cytochrome c oxidase IV
Rn.42785 ESTs, moderately similar to human GP36b glycoprotein
Rn.27371 EST
Rn.2202
CCTACTAACC 77 Rn.1774 Fructose bisphosphate aldolase A
AATAAAAGTT 76 Rn.40255 F1-ATPase alpha subunit
Rn.45103 EST
TCTTTGAACC 75 Rn.10833 ESTs, weakly similar to sarco/endoplasmic reticulum calcium ATPase 1b
TGGAAATGAC 75 Rn.2953 Collagen type I, alpha 1
Rn.8273 Branched chain aminotransferase 1, cytosolic
ATGAAATCAA 74 Rn.29882 ESTs, highly similar to human 40S ribosomal protein S4, X isoform
GCCGAGTGTA 72 Rn.3543 ESTs, highly similar to mouse mitochondrial ATP synthase F chain
CTTGCAAGTG 71 Rn.22045 ESTs, moderately similar to human NADH:ubiquinone oxidoreductase B22 subunit
Rn.34357 ESTs, weakly similar to BAT1
Rn.21831 EST
AATGCCCCCC 71 None Unknown
GATTCCGTGA 70 Rn.5961 Ribosomal protein L37
GCACGGGAAT 70 Rn.34429 Mammalian equivalent of bacterial large ribosomal subunit protein L22
CAGAGTCGCT 68 Rn.7401 ESTs, moderately similar to human ubiquinol-cytochrome c reductase complex 11 kDa precursor
TTGCACAGCC 66 Rn.17580 ESTs, highly similar to tropomyosin 1
ATACTTGACA 65 Rn.25364 ESTs
GCCTAATGTA 65 Rn.2554 Ribosomal protein L21
AATCAACCCG 63 Rn.4229 ESTs, moderately similar to human KIAA0822
TTATGAAATG 63 Rn.3640 ESTs, highly similar to mouse NADH-ubiquinone oxidoreductase MWFE subunit
CCTCACTTTA 62 Rn.40205 ESTs, moderately similar to human ubiquinol-cytochrome c reductase complex 14-kDa protein
GATGCCCCCA 62 None Unknown
TGACTATTAA 62 Rn.32218 EST
Rn.2510 ESTs, weakly similar to mouse ADP-ribosylation factor-like protein 4
(continues)
Table 1B.
 
(continued) Most Frequently Expressed SAGE Tags in EOM
Table 1B.
 
(continued) Most Frequently Expressed SAGE Tags in EOM
Tag Copy No. Unigene Cluster Gene Name, *
GAACATCACT 61 Rn.8483 ESTs, moderately similar to mouse 6.8-kDa mitochondrial proteolipid
GAATCCAACT 60 Rn.3377 ESTs, highly similar to mouse neuronal protein 15.6
AGGAGGCTAC 60 Rn.37556 Ribosomal protein L14
Rn.37667 ESTs
GGTACCGCGG 60 Rn.3549 ESTs
GCATACGGCG 59 Rn.8069 ATP synthase subunit e
CTGCGGCTTC 59 None Unknown
ATGGCATCGT 58 Rn.1318 ESTs, moderately similar to human acyl carrier protein, mitochondrial
AAATCCCGTT 57 None Unknown
AGGCAGACAG 56 Rn.965 Eukaryotic translation elongation factor 1 alpha 2
CCAGAACAGA 56 Rn.36878 Ribosomal protein L30
Rn.42296 EST, moderately similar to 60S ribosomal protein L30
GTGAAGGCGG 56 Rn.11232 Ribosomal protein S3a
CTGGGGCATC 56 None Unknown
AGAGAAGAGT 55 Rn.54672 Myosin heavy chain
Rn.40497 Myosin heavy chain, type IIX
Table 2.
 
Selected Comparisons of Gene Expression Patterns in EOM Versus Leg Muscle
Table 2.
 
Selected Comparisons of Gene Expression Patterns in EOM Versus Leg Muscle
Gene Name EOM SAGE* (%) Skeletal Muscle SAGE (%) Skeletal Muscle EST (%)
Sarcomeric Proteins
 Actin, alpha 1, skeletal muscle 0.2 1.86, ‡ 1.02, ‡
 Myosin heavy chain, type IIX 0.1 0.07 0.45, ‡
 Troponin I, skeletal, fast 2 0.49 0.74, ‡ 0.44
 ESTs, highly similar to tropomyosin 1 0.12 0.27, ‡ 0.35, ‡
 ESTs, highly similar to rat myosin light chain 1, skeletal muscle 0.85 0.47, ‡ 0.37, ‡
 Troponin I, skeletal, slow 1 0.01 0.73, ‡ 0.79, ‡
 Troponin C, slow, † 0.37 0.93, ‡ 0.3
 Troponin T1, slow No tag 0.68 1.25
 Myosin-binding protein C, slow ND 0.21 0.41
 Myosin heavy chain, beta cardiac, slow ND 1.54 1.1
 Myosin light chain 1, ventricular, slow ND 0.55
 Myosin light chain 2, regulatory, cardiac, slow ND 0.36
 Myosin heavy chain 2 (IIA) No tag 1.05 0.54
Carbohydrate/Energy Metabolism
 Glycogen phosphorylase 0.02 0.53, ‡ 0.26, ‡
 Phosphoglycerate mutase ND 0.35
 Creatine kinase, muscle 0.21 1.58, ‡ 2.00, ‡
 Creatine kinase, sarcomeric 0.28 0.04, ‡
 Fructose bisphosphate aldolase A 0.14 1.08, ‡ 0.37, ‡
 Glyceraldehyde-3-phosphate dehydrogenase 0.37 1.10, ‡ 1.20, ‡
 Cytochrome C oxidase (nuclear-encoded isoforms)
  IV 0.14 0.09, §
  Va 0.21 0.06, ‡
  VIa 0.15 0.31, ‡ 0.56, ‡
  VIb 0.18 0.17
  VIc 0.07 0.08 0.12, ¶
  VIIa 0.08 <0.002, ‡ 0.26, ‡
  VIIb 0.39 0.04, ‡ 0.1, ‡
  VIIc 0.19
  VIII 0.32 0.17, ‡
Noncontractile cytoskeleton
 Desmin 0.05 0.74, ‡ 0.35, ‡
 Titin 0.04 0.39, ‡ 0.81, ‡
 Nebulin ND 0.14 0.28
 Vimentin ND 0.11
Table 3.
 
Representation of the Most Frequently Expressed EOM Transcripts in Other Skeletal Muscle Databases
Table 3.
 
Representation of the Most Frequently Expressed EOM Transcripts in Other Skeletal Muscle Databases
Tag Gene EOM SAGE, * (%) Skeletal Muscle SAGE (%) Skeletal Muscle EST (%)
GATGCCCCCC Glu-Pro dipeptide repeat protein 3.85
ATAACACATA NG,NG dimethylarginine dimethylaminohydrolase (Ddah1) 3.27
ATACTGACAC NG,NG dimethylarginine dimethylaminohydrolase (Ddah1) 2.8
AATCGGAGGC CDK110 1.07
AGCCATCCCT Glu-Pro dipeptide repeat protein 0.97
TACCACCTTT ESTs, highly similar to rat myosin light chain 1, skeletal muscle 0.85 0.47, † 0.37, †
TAGGTACAGG NG,NG dimethylarginine dimethylaminohydrolase (Ddah1) 0.71
TTTGGGGAGA Myosin light chain 2, heart 0.66 0.72 0.6
Myosin light chain 2, alkali, ventricular, skeletal, slow
Myosin light chain 2
TGGGTTGTCT Lens epithelial protein 0.66
CTCAGGTCTC Myoglobin 0.59 1.08, † 1.34, †
ATGACTATTA Unknown
GAGGGTCGGA Troponin I, skeletal, fast 2 0.49 0.74, † 0.44
TGGATCCTGA Major beta-globin 0.48 0.03, † 0.48
Zero beta-1 globin, exons 1 and 2
TAGGCCACAC Mitochondrial ATP synthase beta subunit 0.46 0.04, † 0.15, †
AGCGATTCAA ESTs, moderately similar to mouse URF3 0.43 1.3, †
GCCTTCCTCA Unknown
GATAAAACCA ESTs 0.41 0.05, †
Mitochondrial adenine nucleotide translocator
TGGGCACCTG F1-ATPase epsilon subunit 0.4 0.11, †
CCCAGGCACT Troponin T, skeletal, fast 0.39 0.53, † 0.35
GCATATTGAA ESTs, moderately similar to mouse cytochrome c oxidase VIIb precursor 0.39 0.04, † 0.1, †
TGACAGACGA ESTs, weakly similar to human calmodulin 0.37
GCCTCCAAGG Glyceraldehyde-3-phosphate dehydrogenase, brain 0.37 1.1, † 1.2, †
Glyceraldehyde-3-phosphate dehydrogenase
ESTs
CACCCATACC Unknown
CACGCCTCTC Alpha-globin 0.35 0.07, †
Major beta-globin
Hemoglobin, alpha 1
AAAAATCATC ESTs, moderately similar to mouse URF5 0.35 0.35
CTAGTCTTTG Ribosomal protein S29 0.34 0.06, †
AGGAGCTCGG Cytochrome c oxidase subunit VIII-H 0.32 0.17, †
GAAATATGTG ESTs, moderately similar to ATP synthase lipid-binding protein P2 precursor 0.31
TTGTTAGTGC Malate dehydrogenase, cytosolic 0.3 0.04, †
GTGGCTCACA ESTs 0.29
TAAAATTGTA EST, moderately similar to human KIAA0321
ESTs, highly similar to creatine kinase, sarcomeric, mitochondrial, precursor 0.28 0.04, †
GGCTATGTAA ESTs, weakly similar to mouse S43118 finger protein
Decorin 0.25
ESTs
TGTAATGAGA EST 0.23 n
ESTs, moderately similar to mouse F1F0-ATP synthase g subunit 0.05, †
CGAACTCTCA Secreted acidic cysteine-rich glycoprotein (osteonectin) 0.23 0.04, †
AAATAAAACT Tuberous sclerosis 2 0.22
ESTs
Cis-Golgi p28
AGCCGTCCCT Glu-Pro dipeptide repeat protein 0.22
AAAAAAAAAA ESTs 0.21
ATCCCTGCGC Creatine kinase, muscle 0.21 1.58, † 2.0, †
CCAGTCCTGG ATP synthase, H+ transporting, mitochondrial F0 complex, subunit c (subunit 9), isoform 1 0.21 0.08, †
TAACTTGGAC Mitochondrial cytochrome c oxidase subunit Va 0.21 0.06, †
TCAGGCTGCC Ferritin subunit H 0.21 0.11, † 0.28, §
Ferritin
EST, moderately similar to ferritin heavy chain
CTCCTGGACA ESTs, weakly similar to AF099929_1 pervin 0.2
GATCAGTCAT Ribosomal protein L19 0.2 0.1, † 0.19
ESTs
GAATGACCTG Ribosomal protein L13a 0.2 0.34, †
ESTs, highly similar to human 40s ribosomal protein S28 0.11, ‡
AAGATCAAGA Actin, alpha 1, skeletal muscle 0.2 1.86, † 1.02, †
EST
Actin, gamma 2, smooth muscle, enteric 0.86, †
TTGGGCCAGA ESTs, Highly similar to mouse cytochrome c oxidase VIIc precursor 0.19
ESTs
(continues)
Table 3A.
 
Representation of the Most Frequently Expressed EOM Transcripts in Other Skeletal Muscle Databases
Table 3A.
 
Representation of the Most Frequently Expressed EOM Transcripts in Other Skeletal Muscle Databases
Tag Gene EOM SAGE, * (%) Skeletal Muscle SAGE (%) Skeletal Muscle EST (%)
ATGCAACTAC ESTs, highly similar to mouse cytochrome c oxidase VIb 0.18 0.04, † 0.17
CAAACATCCA Glu-Pro dipeptide repeat protein 0.18
GGAGGGGAAG ESTs, moderately similar to mouse URF3 0.18 1.3, †
ESTs
TGTTCCTCTT Procollagen type III, alpha 1 0.18
CGGGATCTGC Oligomycin sensitivity conferring protein 0.18
GGGCAACCAG ESTs, weakly similar to human ubiquinol-cytochrome c reductase complex ubiquinone-binding protein QP-C 0.18 0.21
GGATTTGGCC ESTs, highly similar to acidic ribosomal protein P2 0.18 0.16
GGATTCGGTC Acidic ribosomal protein P0 0.17 0.08, † 0.29, †
TAATACTCAA ESTs 0.17
DAPIT protein
GGCCACCTCC Myosin, heavy polypeptide 13, skeletal muscle 0.17
CCCCATCTCA Parvalbumin 0.17
GAGGCTGTGG ESTs, Weakly similar to Ste20-like kinase 0.17 0.03, †
Phosphoglycerate mutase 0.35, †
TCCAATAAAG ESTs, highly similar to 60S acidic ribosomal protein 0.16 0.22, § 0.45, †
EST
AGCGCCCAGA Cytochrome c oxidase subunit VIa polypeptide 2, heart 0.15 0.31, † 0.56, †
TTAAGACTAG CDK110 0.14
TTAATAAATG Cytochrome c oxidase IV 0.14 0.09, §
ESTs, moderately similar to human GP36b glycoprotein
EST
CCTACTAACC Fructose bisphosphate aldolase A 0.14 1.08, † 0.37, †
AATAAAAGTT F1-ATPase alpha subunit 0.14 0.08, ‡
EST
TCTTTGAACC ESTs, weakly similar to sarco/endoplasmic reticulum calcium ATPase 1b 0.14
TGGAAATGAC Collagen alpha1 type I 0.14
Branched chain aminotransferase 1, cytosolic
ATGAAATCAA ESTs, highly similar to human 40S ribosomal protein S4, X isoform 0.14 0.08, ‡ 0.17
GCCGAGTGTA ESTs, highly similar to mouse mitochondrial ATP synthase F chain 0.13 0.08, §
CTTGCAAGTG ESTs, moderately similar to human NADH:ubiquinone oxidoreductase B22 subunit 0.13
ESTs, weakly similar to BAT1
EST
AATGCCCCCC Unknown
GATTCCGTGA Ribosomal protein L37 0.13 0.08, § 0.57, †
GCACGGGAAT Mammalian equivalent of bacterial large ribosomal subunit protein L22 0.13
CAGAGTCGCT ESTs, moderately similar to human ubiquinol-cytochrome c reductase complex 11-kDa precursor 0.12
TTGCACAGCC ESTs, highly similar to tropomyosin 1 0.12 0.27, † 0.35, †
ATACTTGACA ESTs 0.12
GCCTAATGTA Ribosomal protein L21 0.12 0.11 0.22, †
AATCAACCCG ESTs, moderately similar to human KIAA0822 0.12
TTATGAAATG ESTs, highly similar to mouse NADH-ubiquinone oxidoreductase MWFE subunit 0.12
CCTCACTTTA ESTs, moderately similar to human ubiquinol-cytochrome c reductase complex 14 kDa protein 0.11
GATGCCCCCA Unknown
TGACTATTAA EST ESTs, weakly similar to mouse ADP-ribosylation factor-like protein 4 0.11
GAACATCACT ESTs, moderately similar to mouse 6.8 kDa mitochondrial proteolipid 0.11
GAATCCAACT ESTs, highly similar to mouse neuronal protein 15.6 0.11
AGGAGGCTAC Ribosomal protein L14 0.11 0.11
ESTs
GGTACCGCGG ESTs 0.11
GCATACGGCG ATP synthase subunit e 0.11
CTGCGGCTTC Unknown
ATGGCATCGT ESTs, moderately similar to human acyl carrier protein, mitochondrial 0.11
AAATCCCGTT Unknown
AGGCAGACAG Eukaryotic translation elongation factor 1 alpha 2 0.1 0.19, †
CCAGAACAGA Ribosomal protein L30 0.1 0.08 0.25, †
GTGAAGGCGG Ribosomal protein S3a 0.1 0.05, ‡ 0.47, †
CTGGGGCATC Unknown
AGAGAAGAGT Myosin heavy chain
Myosin heavy chain, type IIX 0.1 0.07 0.45, †
The authors thank Xiaohua Zhou for technical assistance; Beth Ann Benetz for assistance with illustrations; Kenneth V. Kinzler for supplying the SAGE protocol and software; Seth Blackshaw, Connie Cepko, Kornelia Y. Polyak, and Victor Velculescu for advice on microSAGE; Francisco Andrade, Sangeeta Khanna, and Sunil Rao for substantial assistance with data analysis and helpful discussions; and other members of the Porter laboratory for their help. 
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Figure 1.
 
Frequency distribution of EOM-derived SAGE tags. Matched tags denote SAGE tag sequences that map into the NCBI database, and novel tags are those that did not have a sequence match in the database. (A) Semilog plot showing the frequency distribution of SAGE tags detected at specific abundance levels. Novel tags were most frequent among tags expressed only as single copies, and their level declined relative to matched-tag frequency, with increasing copy number. (B) Plot illustrating the frequency of matched SAGE tags. When expressed as a percentage of total tags in each bin, matched-tag frequency increased as a function of copy number.
Figure 1.
 
Frequency distribution of EOM-derived SAGE tags. Matched tags denote SAGE tag sequences that map into the NCBI database, and novel tags are those that did not have a sequence match in the database. (A) Semilog plot showing the frequency distribution of SAGE tags detected at specific abundance levels. Novel tags were most frequent among tags expressed only as single copies, and their level declined relative to matched-tag frequency, with increasing copy number. (B) Plot illustrating the frequency of matched SAGE tags. When expressed as a percentage of total tags in each bin, matched-tag frequency increased as a function of copy number.
Table 1.
 
Most Frequently Expressed SAGE Tags in EOM
Table 1.
 
Most Frequently Expressed SAGE Tags in EOM
Tag Copy No. Unigene Cluster Gene Name, *
GATGCCCCCC 2109 Rn.3979 Glu-Pro dipeptide repeat protein
ATAACACATA 1792 Rn.4241 NG,NG dimethylarginine dimethylaminohydrolase (Ddah1)
ATACTGACAC 1536 Rn.4241 NG,NG dimethylarginine dimethylaminohydrolase (Ddah1)
AATCGGAGGC 584 Rn.25727 CDK110
AGCCATCCCT 530 Rn.3979 Glu-Pro dipeptide repeat protein
TACCACCTTT 464 Rn.40120 ESTs, highly similar to rat myosin light chain 1, skeletal muscle
TAGGTACAGG 387 Rn.4241 NG,NG dimethylarginine dimethylaminohydrolase (Ddah1)
TTTGGGGAGA 364 Rn.37176 Myosin light chain 2, heart
Rn.6534 Myosin light chain 2, alkali, ventricular, skeletal, slow
Rn.54542 Myosin light chain 2
TGGGTTGTCT 361 Rn.36610 Lens epithelial protein
CTCAGGTCTC 322 Rn.40511 Myoglobin
ATGACTATTA 278 None Unknown
GAGGGTCGGA 266 Rn.9924 Troponin I, skeletal, fast 2
TGGATCCTGA 264 Rn.36966 Major beta-globin
Rn.54518 Zero beta-1 globin, exons 1 and 2
TAGGCCACAC 254 Rn.2618 Mitochondrial ATP synthase beta subunit
AGCGATTCAA 236 Rn.43378 ESTs, moderately similar to mouse URF3
GCCTTCCTCA 231 None Unknown
GATAAAACCA 227 Rn.9056 ESTs
Rn.4092 Mitochondrial adenine nucleotide translocator
TGGGCACCTG 220 Rn.3454 F1-ATPase epsilon subunit
CCCAGGCACT 212 Rn.22504 Troponin T, skeletal, fast
GCATATTGAA 211 Rn.2026 ESTs, Moderately similar to mouse cytochrome c oxidase VIIb precursor
TGACAGACGA 203 Rn.43529 ESTs, weakly similar to human calmodulin
GCCTCCAAGG 202 Rn.1925 Glyceraldehyde-3-phosphate dehydrogenase, brain
Rn.54911 Glyceraldehyde-3-phosphate dehydrogenase
Rn.39164 ESTs
CACCCATACC 192 None Unknown
CACGCCTCTC 191 Rn.55177 Alpha-globin
Rn.36966 Major beta-globin
Rn.11229 Hemoglobin, alpha 1
AAAAATCATC 190 Rn.7381 ESTs, moderately similar to mouse URF5
CTAGTCTTTG 186 Rn.34942 Ribosomal protein S29
AGGAGCTCGG 177 Rn.10325 Cytochrome c oxidase subunit VIII-H
GAAATATGTG 168 Rn.2180 ESTs, moderately similar to ATP synthase lipid-binding protein P2 precursor
TTGTTAGTGC 167 Rn.1504 Malate dehydrogenase, cytosolic
GTGGCTCACA 161 Rn.51650 ESTs
Rn.40562
Rn.53514
Rn.51174
Rn.44604
Rn.43236
Rn.52705
Rn.40249
Rn.42743
Rn.43716
TAAAATTGTA 152 Rn.32408 EST, moderately similar to human KIAA0321
Rn.3905 ESTs, highly similar to creatine kinase, sarcomeric, mitochondrial, precursor
GGCTATGTAA 136 Rn.51390 ESTs, weakly similar to mouse S43118 finger protein
Rn.3819 Decorin
Rn.20597 ESTs
TGTAATGAGA 128 Rn.52850 EST
Rn.2462 ESTs, moderately similar to mouse F1F0-ATP synthase g subunit
CGAACTCTCA 124 Rn.31991 Secreted acidic cysteine-rich glycoprotein (osteonectin)
AAATAAAACT 123 Rn.5875 Tuberous sclerosis 2
Rn.40213 ESTs
Rn.6390 Cis-Golgi p28
Rn.27383
AGCCGTCCCT 120 Rn.3979 Glu-Pro dipeptide repeat protein
AAAAAAAAAA 116 Rn.20806 ESTs
Rn.20483
Rn.45252
Rn.10764
Rn.11656
Rn.43473
Rn.11739
Rn.14962
Rn.14882
Rn.2122
(continues)
Table 1A.
 
(continued) Most Frequently Expressed SAGE Tags in EOM
Table 1A.
 
(continued) Most Frequently Expressed SAGE Tags in EOM
Tag Copy No. Unigene Cluster Gene Name, *
ATCCCTGCGC 116 Rn.10756 Creatine kinase, muscle
CCAGTCCTGG 116 Rn.3357 ATP synthase, H+ transporting, mitochondrial F0 complex, subunit c (subunit 9), isoform 1
TAACTTGGAC 115 Rn.11077 Mitochondrial cytochrome c oxidase subunit Va
TCAGGCTGCC 114 Rn.1960 Ferritin subunit H
Rn.54798 Ferritin
Rn.42678 EST, moderately similar to ferritin heavy chain
CTCCTGGACA 111 Rn.2883 ESTs, weakly similar to AF099929_1 pervin
GATCAGTCAT 108 Rn.3384 Ribosomal protein L19
Rn.40331 ESTs
GAATGACCTG 108 Rn.34334 Ribosomal protein L13a
Rn.6005 ESTs, highly similar to human 40s ribosomal protein S28
AAGATCAAGA 107 Rn.39438 Actin, alpha 1, skeletal muscle
Rn.50809 EST
Rn.958 Actin, gamma 2, smooth muscle, enteric
Rn.69
TTGGGCCAGA 104 Rn.2130 ESTs, Highly similar to mouse cytochrome c oxidase VIIc precursor
Rn.21453 ESTs
ATGCAACTAC 101 Rn.6009 ESTs, highly similar to mouse cytochrome c oxidase VIb
CAAACATCCA 99 Rn.3979 Glu-Pro dipeptide repeat protein
GGAGGGGAAG 98 Rn.43378 ESTs, moderately similar to mouse URF3
Rn.22772 ESTs
TGTTCCTCTT 97 Rn.3247 Procollagen type III, alpha 1
CGGGATCTGC 97 Rn.1817 Oligomycin sensitivity conferring protein
GGGCAACCAG 97 Rn.3472 ESTs, weakly similar to human ubiquinol-cytochrome c reductase complex ubiquinone-binding protein QP-C
GGATTTGGCC 96 Rn.55312 ESTs, highly similar to acidic ribosomal protein P2
Rn.976 ESTs, highly similar to 60S acidic ribosomal protein P2
GGATTCGGTC 94 Rn.1079 Acidic ribosomal protein P0
TAATACTCAA 94 Rn.7661 ESTs
Rn.4281 DAPIT protein
GGCCACCTCC 93 Rn.34319 Myosin, heavy polypeptide 13, skeletal muscle
CCCCATCTCA 92 Rn.2005 Parvalbumin
GAGGCTGTGG 91 Rn.12939 ESTs, weakly similar to Ste20-like kinase
Rn.9738 Phosphoglycerate mutase
TCCAATAAAG 87 Rn.973 ESTs, highly similar to 60S acidic ribosomal protein
Rn.47395 EST
AGCGCCCAGA 83 Rn.5119 Cytochrome c oxidase subunit VIa polypeptide 2, heart
TTAAGACTAG 78 Rn.25727 CDK110
TTAATAAATG 78 Rn.2528 Cytochrome c oxidase IV
Rn.42785 ESTs, moderately similar to human GP36b glycoprotein
Rn.27371 EST
Rn.2202
CCTACTAACC 77 Rn.1774 Fructose bisphosphate aldolase A
AATAAAAGTT 76 Rn.40255 F1-ATPase alpha subunit
Rn.45103 EST
TCTTTGAACC 75 Rn.10833 ESTs, weakly similar to sarco/endoplasmic reticulum calcium ATPase 1b
TGGAAATGAC 75 Rn.2953 Collagen type I, alpha 1
Rn.8273 Branched chain aminotransferase 1, cytosolic
ATGAAATCAA 74 Rn.29882 ESTs, highly similar to human 40S ribosomal protein S4, X isoform
GCCGAGTGTA 72 Rn.3543 ESTs, highly similar to mouse mitochondrial ATP synthase F chain
CTTGCAAGTG 71 Rn.22045 ESTs, moderately similar to human NADH:ubiquinone oxidoreductase B22 subunit
Rn.34357 ESTs, weakly similar to BAT1
Rn.21831 EST
AATGCCCCCC 71 None Unknown
GATTCCGTGA 70 Rn.5961 Ribosomal protein L37
GCACGGGAAT 70 Rn.34429 Mammalian equivalent of bacterial large ribosomal subunit protein L22
CAGAGTCGCT 68 Rn.7401 ESTs, moderately similar to human ubiquinol-cytochrome c reductase complex 11 kDa precursor
TTGCACAGCC 66 Rn.17580 ESTs, highly similar to tropomyosin 1
ATACTTGACA 65 Rn.25364 ESTs
GCCTAATGTA 65 Rn.2554 Ribosomal protein L21
AATCAACCCG 63 Rn.4229 ESTs, moderately similar to human KIAA0822
TTATGAAATG 63 Rn.3640 ESTs, highly similar to mouse NADH-ubiquinone oxidoreductase MWFE subunit
CCTCACTTTA 62 Rn.40205 ESTs, moderately similar to human ubiquinol-cytochrome c reductase complex 14-kDa protein
GATGCCCCCA 62 None Unknown
TGACTATTAA 62 Rn.32218 EST
Rn.2510 ESTs, weakly similar to mouse ADP-ribosylation factor-like protein 4
(continues)
Table 1B.
 
(continued) Most Frequently Expressed SAGE Tags in EOM
Table 1B.
 
(continued) Most Frequently Expressed SAGE Tags in EOM
Tag Copy No. Unigene Cluster Gene Name, *
GAACATCACT 61 Rn.8483 ESTs, moderately similar to mouse 6.8-kDa mitochondrial proteolipid
GAATCCAACT 60 Rn.3377 ESTs, highly similar to mouse neuronal protein 15.6
AGGAGGCTAC 60 Rn.37556 Ribosomal protein L14
Rn.37667 ESTs
GGTACCGCGG 60 Rn.3549 ESTs
GCATACGGCG 59 Rn.8069 ATP synthase subunit e
CTGCGGCTTC 59 None Unknown
ATGGCATCGT 58 Rn.1318 ESTs, moderately similar to human acyl carrier protein, mitochondrial
AAATCCCGTT 57 None Unknown
AGGCAGACAG 56 Rn.965 Eukaryotic translation elongation factor 1 alpha 2
CCAGAACAGA 56 Rn.36878 Ribosomal protein L30
Rn.42296 EST, moderately similar to 60S ribosomal protein L30
GTGAAGGCGG 56 Rn.11232 Ribosomal protein S3a
CTGGGGCATC 56 None Unknown
AGAGAAGAGT 55 Rn.54672 Myosin heavy chain
Rn.40497 Myosin heavy chain, type IIX
Table 2.
 
Selected Comparisons of Gene Expression Patterns in EOM Versus Leg Muscle
Table 2.
 
Selected Comparisons of Gene Expression Patterns in EOM Versus Leg Muscle
Gene Name EOM SAGE* (%) Skeletal Muscle SAGE (%) Skeletal Muscle EST (%)
Sarcomeric Proteins
 Actin, alpha 1, skeletal muscle 0.2 1.86, ‡ 1.02, ‡
 Myosin heavy chain, type IIX 0.1 0.07 0.45, ‡
 Troponin I, skeletal, fast 2 0.49 0.74, ‡ 0.44
 ESTs, highly similar to tropomyosin 1 0.12 0.27, ‡ 0.35, ‡
 ESTs, highly similar to rat myosin light chain 1, skeletal muscle 0.85 0.47, ‡ 0.37, ‡
 Troponin I, skeletal, slow 1 0.01 0.73, ‡ 0.79, ‡
 Troponin C, slow, † 0.37 0.93, ‡ 0.3
 Troponin T1, slow No tag 0.68 1.25
 Myosin-binding protein C, slow ND 0.21 0.41
 Myosin heavy chain, beta cardiac, slow ND 1.54 1.1
 Myosin light chain 1, ventricular, slow ND 0.55
 Myosin light chain 2, regulatory, cardiac, slow ND 0.36
 Myosin heavy chain 2 (IIA) No tag 1.05 0.54
Carbohydrate/Energy Metabolism
 Glycogen phosphorylase 0.02 0.53, ‡ 0.26, ‡
 Phosphoglycerate mutase ND 0.35
 Creatine kinase, muscle 0.21 1.58, ‡ 2.00, ‡
 Creatine kinase, sarcomeric 0.28 0.04, ‡
 Fructose bisphosphate aldolase A 0.14 1.08, ‡ 0.37, ‡
 Glyceraldehyde-3-phosphate dehydrogenase 0.37 1.10, ‡ 1.20, ‡
 Cytochrome C oxidase (nuclear-encoded isoforms)
  IV 0.14 0.09, §
  Va 0.21 0.06, ‡
  VIa 0.15 0.31, ‡ 0.56, ‡
  VIb 0.18 0.17
  VIc 0.07 0.08 0.12, ¶
  VIIa 0.08 <0.002, ‡ 0.26, ‡
  VIIb 0.39 0.04, ‡ 0.1, ‡
  VIIc 0.19
  VIII 0.32 0.17, ‡
Noncontractile cytoskeleton
 Desmin 0.05 0.74, ‡ 0.35, ‡
 Titin 0.04 0.39, ‡ 0.81, ‡
 Nebulin ND 0.14 0.28
 Vimentin ND 0.11
Table 3.
 
Representation of the Most Frequently Expressed EOM Transcripts in Other Skeletal Muscle Databases
Table 3.
 
Representation of the Most Frequently Expressed EOM Transcripts in Other Skeletal Muscle Databases
Tag Gene EOM SAGE, * (%) Skeletal Muscle SAGE (%) Skeletal Muscle EST (%)
GATGCCCCCC Glu-Pro dipeptide repeat protein 3.85
ATAACACATA NG,NG dimethylarginine dimethylaminohydrolase (Ddah1) 3.27
ATACTGACAC NG,NG dimethylarginine dimethylaminohydrolase (Ddah1) 2.8
AATCGGAGGC CDK110 1.07
AGCCATCCCT Glu-Pro dipeptide repeat protein 0.97
TACCACCTTT ESTs, highly similar to rat myosin light chain 1, skeletal muscle 0.85 0.47, † 0.37, †
TAGGTACAGG NG,NG dimethylarginine dimethylaminohydrolase (Ddah1) 0.71
TTTGGGGAGA Myosin light chain 2, heart 0.66 0.72 0.6
Myosin light chain 2, alkali, ventricular, skeletal, slow
Myosin light chain 2
TGGGTTGTCT Lens epithelial protein 0.66
CTCAGGTCTC Myoglobin 0.59 1.08, † 1.34, †
ATGACTATTA Unknown
GAGGGTCGGA Troponin I, skeletal, fast 2 0.49 0.74, † 0.44
TGGATCCTGA Major beta-globin 0.48 0.03, † 0.48
Zero beta-1 globin, exons 1 and 2
TAGGCCACAC Mitochondrial ATP synthase beta subunit 0.46 0.04, † 0.15, †
AGCGATTCAA ESTs, moderately similar to mouse URF3 0.43 1.3, †
GCCTTCCTCA Unknown
GATAAAACCA ESTs 0.41 0.05, †
Mitochondrial adenine nucleotide translocator
TGGGCACCTG F1-ATPase epsilon subunit 0.4 0.11, †
CCCAGGCACT Troponin T, skeletal, fast 0.39 0.53, † 0.35
GCATATTGAA ESTs, moderately similar to mouse cytochrome c oxidase VIIb precursor 0.39 0.04, † 0.1, †
TGACAGACGA ESTs, weakly similar to human calmodulin 0.37
GCCTCCAAGG Glyceraldehyde-3-phosphate dehydrogenase, brain 0.37 1.1, † 1.2, †
Glyceraldehyde-3-phosphate dehydrogenase
ESTs
CACCCATACC Unknown
CACGCCTCTC Alpha-globin 0.35 0.07, †
Major beta-globin
Hemoglobin, alpha 1
AAAAATCATC ESTs, moderately similar to mouse URF5 0.35 0.35
CTAGTCTTTG Ribosomal protein S29 0.34 0.06, †
AGGAGCTCGG Cytochrome c oxidase subunit VIII-H 0.32 0.17, †
GAAATATGTG ESTs, moderately similar to ATP synthase lipid-binding protein P2 precursor 0.31
TTGTTAGTGC Malate dehydrogenase, cytosolic 0.3 0.04, †
GTGGCTCACA ESTs 0.29
TAAAATTGTA EST, moderately similar to human KIAA0321
ESTs, highly similar to creatine kinase, sarcomeric, mitochondrial, precursor 0.28 0.04, †
GGCTATGTAA ESTs, weakly similar to mouse S43118 finger protein
Decorin 0.25
ESTs
TGTAATGAGA EST 0.23 n
ESTs, moderately similar to mouse F1F0-ATP synthase g subunit 0.05, †
CGAACTCTCA Secreted acidic cysteine-rich glycoprotein (osteonectin) 0.23 0.04, †
AAATAAAACT Tuberous sclerosis 2 0.22
ESTs
Cis-Golgi p28
AGCCGTCCCT Glu-Pro dipeptide repeat protein 0.22
AAAAAAAAAA ESTs 0.21
ATCCCTGCGC Creatine kinase, muscle 0.21 1.58, † 2.0, †
CCAGTCCTGG ATP synthase, H+ transporting, mitochondrial F0 complex, subunit c (subunit 9), isoform 1 0.21 0.08, †
TAACTTGGAC Mitochondrial cytochrome c oxidase subunit Va 0.21 0.06, †
TCAGGCTGCC Ferritin subunit H 0.21 0.11, † 0.28, §
Ferritin
EST, moderately similar to ferritin heavy chain
CTCCTGGACA ESTs, weakly similar to AF099929_1 pervin 0.2
GATCAGTCAT Ribosomal protein L19 0.2 0.1, † 0.19
ESTs
GAATGACCTG Ribosomal protein L13a 0.2 0.34, †
ESTs, highly similar to human 40s ribosomal protein S28 0.11, ‡
AAGATCAAGA Actin, alpha 1, skeletal muscle 0.2 1.86, † 1.02, †
EST
Actin, gamma 2, smooth muscle, enteric 0.86, †
TTGGGCCAGA ESTs, Highly similar to mouse cytochrome c oxidase VIIc precursor 0.19
ESTs
(continues)
Table 3A.
 
Representation of the Most Frequently Expressed EOM Transcripts in Other Skeletal Muscle Databases
Table 3A.
 
Representation of the Most Frequently Expressed EOM Transcripts in Other Skeletal Muscle Databases
Tag Gene EOM SAGE, * (%) Skeletal Muscle SAGE (%) Skeletal Muscle EST (%)
ATGCAACTAC ESTs, highly similar to mouse cytochrome c oxidase VIb 0.18 0.04, † 0.17
CAAACATCCA Glu-Pro dipeptide repeat protein 0.18
GGAGGGGAAG ESTs, moderately similar to mouse URF3 0.18 1.3, †
ESTs
TGTTCCTCTT Procollagen type III, alpha 1 0.18
CGGGATCTGC Oligomycin sensitivity conferring protein 0.18
GGGCAACCAG ESTs, weakly similar to human ubiquinol-cytochrome c reductase complex ubiquinone-binding protein QP-C 0.18 0.21
GGATTTGGCC ESTs, highly similar to acidic ribosomal protein P2 0.18 0.16
GGATTCGGTC Acidic ribosomal protein P0 0.17 0.08, † 0.29, †
TAATACTCAA ESTs 0.17
DAPIT protein
GGCCACCTCC Myosin, heavy polypeptide 13, skeletal muscle 0.17
CCCCATCTCA Parvalbumin 0.17
GAGGCTGTGG ESTs, Weakly similar to Ste20-like kinase 0.17 0.03, †
Phosphoglycerate mutase 0.35, †
TCCAATAAAG ESTs, highly similar to 60S acidic ribosomal protein 0.16 0.22, § 0.45, †
EST
AGCGCCCAGA Cytochrome c oxidase subunit VIa polypeptide 2, heart 0.15 0.31, † 0.56, †
TTAAGACTAG CDK110 0.14
TTAATAAATG Cytochrome c oxidase IV 0.14 0.09, §
ESTs, moderately similar to human GP36b glycoprotein
EST
CCTACTAACC Fructose bisphosphate aldolase A 0.14 1.08, † 0.37, †
AATAAAAGTT F1-ATPase alpha subunit 0.14 0.08, ‡
EST
TCTTTGAACC ESTs, weakly similar to sarco/endoplasmic reticulum calcium ATPase 1b 0.14
TGGAAATGAC Collagen alpha1 type I 0.14
Branched chain aminotransferase 1, cytosolic
ATGAAATCAA ESTs, highly similar to human 40S ribosomal protein S4, X isoform 0.14 0.08, ‡ 0.17
GCCGAGTGTA ESTs, highly similar to mouse mitochondrial ATP synthase F chain 0.13 0.08, §
CTTGCAAGTG ESTs, moderately similar to human NADH:ubiquinone oxidoreductase B22 subunit 0.13
ESTs, weakly similar to BAT1
EST
AATGCCCCCC Unknown
GATTCCGTGA Ribosomal protein L37 0.13 0.08, § 0.57, †
GCACGGGAAT Mammalian equivalent of bacterial large ribosomal subunit protein L22 0.13
CAGAGTCGCT ESTs, moderately similar to human ubiquinol-cytochrome c reductase complex 11-kDa precursor 0.12
TTGCACAGCC ESTs, highly similar to tropomyosin 1 0.12 0.27, † 0.35, †
ATACTTGACA ESTs 0.12
GCCTAATGTA Ribosomal protein L21 0.12 0.11 0.22, †
AATCAACCCG ESTs, moderately similar to human KIAA0822 0.12
TTATGAAATG ESTs, highly similar to mouse NADH-ubiquinone oxidoreductase MWFE subunit 0.12
CCTCACTTTA ESTs, moderately similar to human ubiquinol-cytochrome c reductase complex 14 kDa protein 0.11
GATGCCCCCA Unknown
TGACTATTAA EST ESTs, weakly similar to mouse ADP-ribosylation factor-like protein 4 0.11
GAACATCACT ESTs, moderately similar to mouse 6.8 kDa mitochondrial proteolipid 0.11
GAATCCAACT ESTs, highly similar to mouse neuronal protein 15.6 0.11
AGGAGGCTAC Ribosomal protein L14 0.11 0.11
ESTs
GGTACCGCGG ESTs 0.11
GCATACGGCG ATP synthase subunit e 0.11
CTGCGGCTTC Unknown
ATGGCATCGT ESTs, moderately similar to human acyl carrier protein, mitochondrial 0.11
AAATCCCGTT Unknown
AGGCAGACAG Eukaryotic translation elongation factor 1 alpha 2 0.1 0.19, †
CCAGAACAGA Ribosomal protein L30 0.1 0.08 0.25, †
GTGAAGGCGG Ribosomal protein S3a 0.1 0.05, ‡ 0.47, †
CTGGGGCATC Unknown
AGAGAAGAGT Myosin heavy chain
Myosin heavy chain, type IIX 0.1 0.07 0.45, †
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