Broader coverage of gene bodies with LNA/RNA-hybrid template-switching oligonucleotides
We have compared template-switching oligonucleotides of different chemical nature, namely the standard DNA/RNA hybrids with DNA/LNA and pure DNA alternatives, hoping to increase the efficiency of the nanoCAGE protocol. To our surprise the repartition of the signal obtained with the DNA/LNA and the pure DNA oligonucleotides covered gene bodies more flatly, that is, did not show the same preference for the 5′ end as with DNA/RNA.
We published these results in BMC Genomics, where we discussed the implications for the mechanism of template switching, and the possible uses of the DNA/LNA oligonucleotides. This article also features an extensive source code listing in its supplementary material, which may be useful to people doing pairwise comparisons of triplicated CAGE libraries.
While our article was in revision, another study of LNA-containing oligonucleotides (a DNA/RNA/LNA triple hybrid) was published by Picelli et al.
Harbers M, Kato S, de Hoon M, Hayashizaki Y, Carninci P, Plessy C. Comparison of RNA- or LNA-hybrid oligonucleotides in template-switching reactions for high-speed sequencing library preparation. BMC Genomics. 2013 Sep 30 14(1):665. PubMed: 24079827