Equalization of Gene Representation
Expression of genes varies from a few to
several thousand copies (mRNA molecules) in certain cell types. This
high range of difference in gene representation makes it
difficult to discover rare mRNAs even with the new ultra-high-throughput
sequencing technologies, especially when using PacBio RS II sequencing. To
redundancy in cDNA populations, the representation of each sequence has
VERTIS offers a well-proven and highly efficient cDNA normalization using a kinetic
denaturation-reassociation technique as outlined in the schema below. Already after
one round of normalization nearly equal representation of each gene is
achieved and a typical
normalized cDNA library has a greater than 100-fold reduction in highly
abundant clones, such as beta-actin.
Normalized cDNA libraries are also very useful for sequencing with the Illumina MiSeq system. The MiSeq allows read lengths of up to 2x300 bp, but the data output is restricted to about 25 million reads per run. Therefore, the use of cDNA libraries with reduced redundancy can substantially increase sequence coverage per MiSeq sequencing run.
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Pauli, Martina, et al. "De novo assembly of the dual transcriptomes of a polymorphic raptor species and its malarial parasite." BMC genomics 16.1 (2015): 1.
Lin, Ya-Fen, et al. "A comprehensive set of transcript sequences of the heavy metal hyperaccumulator Noccaea caerulescens." Front Plant Sci 5 (2014): 261.
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