Design Of The Rnai Vector Essay

1625 Words Feb 10th, 2015 null Page
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Design of the RNAi vector
The PMT gene family is well conserved among the Solanales. There are five genes in the PMT gene family (PMT1 to PMT5) which differ in the exon 1 sequence due to the insertion of a variable number of 33 nucleotide repeats. This makes the first exon as an ideal region for the design of gene specific RNAi trigger sequence (Fig 1A). Full-length sequences are available for four PMT genes (PMT1 to PMT4) whereas only the sequence of exon1 is known for PMT5 which shares 95% identity with PMT1. The PMT4 contains 6 insertions of 33 nucleotide repeats (198 bases) in exon 1. The exon 1 sequences of the five PMT genes were analyzed to design a trigger sequence that is specific to the major PMT gene, PMT2. A 162bp sequence from the first exon of PMT2 gene was selected as the RNAi trigger sequence that could be more specific to this gene. The 162bp RNAi trigger sequence shares a stretch of 22 nucleotides with PMT3 gene and two stretches of 22 nucleotides each with PMT1 and PMT5 genes. PMT4 gene shares minimum three 25 nucleotides long stretch with the RNAi trigger (Fig 1B). Therefore, the design of the trigger though targets PMT2 gene; it may also silence PMT4 gene. Prediction analysis using RNAi design tools shows the presence of four potential guide siRNAs in the 162bp RNAi trigger sequence. The guide sequences have delta G ranges from -24.21 to -34.51 kcal/Mol and GC content 32 to 53%. The nucleotide mismatches between the predicted guide siRNA and PMT…

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