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Richard W. McKenzie, James E. Jumblatt, Marcia M. Jumblatt; Quantification of MUC2 and MUC5AC Transcripts in Human Conjunctiva. Invest. Ophthalmol. Vis. Sci. 2000;41(3):703-708.
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© ARVO (1962-2015); The Authors (2016-present)
purpose. Transcripts of mucins 1, 4, and 5AC have been identified in human
conjunctival tissue. Of these, only MUC5AC has been localized to goblet
cells. MUC2 is a goblet cell mucin originally identified in the
intestinal mucosa. The presence of MUC2 transcripts and levels of MUC2
and MUC5AC transcripts in normal human conjunctiva, as determined by
quantitative polymerase chain reaction (PCR), is reported.
methods. RNA from conjunctivae of six donors (3 men, 3 women, 44 to 69 years,
all white) was isolated and subjected to competitive reverse
transcription–PCR. Internal standards, which are dsDNA molecules with
ends complementary to a given primer pair but containing nonhomologous
central sequences, were prepared for each gene assayed. Titration of a
constant amount of cDNA against serial dilutions of the internal
standard allowed quantification of the template cDNA. MUC2 and MUC5AC
levels were compared to levels of the “housekeeping” gene,β 2-microglobulin (β2M). The identity of PCR
products was confirmed by sequencing.
results. In the six individual samples tested, β2M mRNA is
expressed, on average, at approximately 10−20 moles per
sample (1 μg RNA) or approximately 63.5 × 104 molecules. The mRNA encoding MUC5AC, a relatively abundant ocular
mucin, exists at levels 10-fold lower than β2M. In
contrast to previous reports of MUC2 mRNA being absent at the ocular
surface, these results show that MUC2 transcripts are present and
expressed at levels 5900-fold lower than for MUC5AC. Apparently, MUC2
transcripts exist on the order of only approximately 100 to 1000
molecules/μg of RNA in the analyzed samples.
conclusions. MUC2 transcripts are present in human conjunctival tissue and their
abundance is much lower than that of MUC5AC. This is the first
application of competitive PCR to the quantitative analysis of mucin
expression in human ocular tissue. The sensitivity of this method
allows the detection of MUC2 transcripts that were not detected by
Northern blot analysis or in situ hybridization in previous studies. It
also makes possible the comparison of relative levels of expression for
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