Splicist™ Library Panels
For discovery of novel transcripts by PCR
While nearly all multi-exon genes are alternatively spliced, many splice isoforms expressed in normal
tissue remain unknown. Moreover, the set of altered transcripts responsible for oncogenesis, whether
aberrantly spliced or otherwise mutated, remain incompletely characterized for important cancers.
- Broad coverage of the transcriptome in 96-well format
- Nearly 5 million clones represented per panel
- Enriched for full-length while preserving the diversity of messages
- Low-complexity format enables discovery of rare transcripts
- PCR primers easily designed with the RefSplice utility
Now for the first time, broad samples of the mammalian polyA+ transcriptome are available in single
96-well microplates. Containing cDNAs derived from normal tissues or cancer cell lines, these panels
provide an efficient way to quickly survey the human transcriptome for novel transcripts. Genofi's
innovative cloning technique minimizes artifacts like chimeras and genomic
contamination, while the separate handling of cDNA size fractions preserves
the diversity of messages found in the original source material. Together these procedures enrich the
resulting libraries for full-length transcripts.
Because individual cDNAs are distributed into low-complexity pools of 50,000 clones per well,
the Splicist panels enable discovery of rare transcripts, while distinguishing highly similar
transcripts. This is so because of the enhanced probability that a particular transcript will be
the sole representative of its parent gene in a pool. PCR performed on a more traditional high-complexity
library pool (>1 million clones) would enable detection of only the most abundant transcripts,
while the rarer transcripts would go undetected.
Screening the Splicist Panel is a simple process. Using the RefSplice utility, it is easy to design PCR primers to hybridize with any pair of
exons in order to amplify all intervening cDNA sequences present in some pools. PCR conditions
are first optimized using a Splicist Tester, which contains the entire library complexity of the
corresponding panel in a single tube. After optimization, PCR is performed on the 96-well
microplate and the resulting amplicons evaluated by gel electrophoresis (see, for example,
results for Hs. IL4R) followed by direct DNA sequencing to discover novel transcripts.
These newly identified sequences, in turn, enable the design of transcript-specific primers for
the further interrogation of specific tissues and cell lines or can be subcloned for functional
analysis. Glycerol stocks for any of the positive pools can be ordered to obtain more material
for further evaluation or to retrieve the original clone using conventional colony screening methods.
While a Splicist Tester is provided with each library panel kit to optimize PCR conditions before
screening the 96-well plates, it is also available separately to determine whether your gene of
interest is present in a specific Splicist panel.
|Splicist Human Universal Panel
|Kit consists of four identical 96-well plates containing
the Splicist cDNA Library template (dried), a Splicist Human Universal Tester
(cat no. SHUT-010) to optimize PCR conditions, and a control primer set.
|Splicist Human Brain Panel
|Kit consists of four identical 96-well plates containing the Splicist
Brain cDNA template (dried), a Splicist Human Brain Tester (cat no. SHBT-010) to
optimize PCR conditions, and a control primer set.
1. Wang, E.T. et al. Alternative isoform regulation in human tissue transcriptomes. Nature 456, 470-6 (2008)
2. Venables, J.P. et al. Cancer-associated regulation of alternative splicing. Nat Struct Mol Biol. 16, 670-6 (2009)