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Adapting 3' Rapid Amplification of CDNA Ends to Map Transcripts in Cancer
Published on: March 28, 2018
Next-generation sequencing-based 5' rapid amplification of cDNA ends for alternative promoters
Bambarendage P U Perera1, Joomyeong Kim1
1Department of Biological Sciences, Louisiana State University, Baton Rouge, LA 70803, USA.
Abstract:
Mammalian genomes contain many unknown alternative first exons and promoters. Thus, we have modified the existing 5'RACE (5' rapid amplification of cDNA ends) approach into a next-generation sequencing (NGS)-based new protocol that can identify these alternative promoters. This protocol has incorporated two main ideas: (i) 5'RACE starting from the known second exons of genes and (ii) NGS-based sequencing of the subsequent cDNA products. This protocol also provides a bioinformatics strategy that processes the sequence reads from NGS runs. This protocol has successfully identified several alternative promoters for an imprinted gene, PEG3. Overall, this NGS-based 5'RACE protocol is a sensitive and reliable method for detecting low-abundant transcripts and promoters.
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