Abstract
Next-generation sequencing (NGS), particularly RNA-sequencing (RNA-Seq) technique, allows detection and quantification of different RNA transcripts in a tissue sample, and in our case toxin transcripts from snake venom glands. Using this approach, novel toxin transcripts can be detected and abundancies of different isoforms of each toxin measured. The analytical pipeline can be briefly outlined as follows. Isolation of mRNA from tissue under RNase-free condition is essential to keep mRNA intact before sequencing. After mRNA fragmentation, the adapters are added to both ends of the fragments to synthesize complementary cDNAs. The obtained cDNA library is then sequenced on Illumina HiSeq 2000 platform. Quality of millions of reads produced from the NGS is checked and the sequences corresponding to the adapters and low-quality reads are removed. Subsequently, the NGS data are subjected to the workflow of de novo assembly, quantification of expression levels, annotation of transcripts, and identification of ORFs, signal peptides, structurally conserved domains, and functional motifs. In this report we describe the listed methodological steps and techniques in details and refer to the platforms and software that may be adopted for similar studies.
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Acknowledgment
We would like to thank Dr. Lawan Chanhome, Head of the Snake Farm, Queen Saovabha Memorial Institute, The Thai Red Cross Society, Thailand; Officers and Staffs from the Snake Farm, Myanmar Pharmaceutical Factory, Yangon, Myanmar for their kind help in venom gland extraction.
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Yee, K.T., Vasieva, O., Rojnuckarin, P. (2020). RNA-Sequencing of Snake Venom Glands. In: Priel, A. (eds) Snake and Spider Toxins. Methods in Molecular Biology, vol 2068. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-9845-6_5
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DOI: https://doi.org/10.1007/978-1-4939-9845-6_5
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Publisher Name: Humana, New York, NY
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Online ISBN: 978-1-4939-9845-6
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