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cross-species array platforms (Table
4.1
), the major factor contributing to the lack
of CHO specific miRNA expression data—the absence in public miRNA sequence
data—has only recently been overcome.
4.3
Breaking the Ground for CHO-Specific miRNA Profiling
The often cited lack of availability of public information on genomic and tran-
scriptomic sequences derived from CHO cells has resulted in an expanded use of
cross-species platforms for measuring RNA transcript levels. While this requires
sophisticated verification of results for mRNA array platforms (Yee et al.
2008
;
Ernst et al.
2006
), the finding that miRNA sequences are generally well conserved
across animals, with an even higher degree of conservation between human, mouse
and rat (Berezikov
2005
), has simplified this task for cross-species miRNA plat-
forms. Consequently, all currently available CHO miRNA profiling data (Table
4.1
)
were acquired using either human, mouse or rat platforms except for two studies,
which applied next-generation sequencing (RNA-seq) technologies (Hackl et al.
2011
; Johnson et al.
2011
).
Sequence data collected in these studies have been included in publications,
and are likely to be incorporated into the miRBase miRNA sequence repository—
a database harboring all currently known mature as well as precursor miRNA
sequences—during its next update (Griffiths-Jones
2010
). Since these studies were
not primarily designed to pinpoint differentially expressed miRNAs, but to identify
and annotate the maximum number of CHO miRNA sequences, a variety of different
cell types and culture conditions were included in the sequencing efforts: sequenc-
ing libraries were generated from distinct CHO cell lines representing (i) adherent
and suspension adapted cells, (ii) host and recombinant cells, (iii) cells cultivated in
serum-free and serum-containing media and (iv) from a pool of RNA harvested from
growth arrested CHO cells or after cold shock, heat shock or sodium butyrate treat-
ment. By BLAST alignment or mapping sequencing reads to a reference composed
of miRBase miRNA sequences, a high degree of miRNA sequence conservation
between human, mouse, rat and Chinese hamster (
Cricetulus grisesus
, abbreviated
with
cgr
in miRNA nomenclature) was confirmed: 350 conserved miRNAs (Johnson
et al.
2011
) compared to 380 conserved miRNAs (Hackl et al.
2011
) were reported,
which overlap by 340 miRNA sequences and therefore add up to a total of 390 cur-
rently identified mature miRNAs in CHO cells. Although the numbers of mature
miRNAs reported for human (1,921), mouse (1,157) or rat (680) in miRBase version
18 are higher, tissue and cell type specific expression of miRNAs results in much
lower numbers of expressed miRNAs in a cell line (Lee et al.
2008
). Therefore, it
is likely the set of 390 mature miRNAs reported in CHO constitutes the majority of
highly conserved miRNAs that are expressed in CHO cells.
Based on the publication of the CHO-K1 genome in August 2011 (Xu et al.
2011
),
we also started to look at the genomic location of these 390 CHO miRNAs in the
given reference genome (Hackl et al.
2012
): using BLAST alignment unambiguous
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