Polygene interferential shRNA plasmid expression vector, construction method and application

Abstract

The invention discloses a plasmid vector pSilencer-U6 / H1-(shRNA)n which can be used for simultaneously expressing a plurality of shRNA, a construction method and application thereof. The vector is composed of two groups of sequences of multiple cloning sites (MCS), a plurality of target gene shRNA transcription units and plasmid sequences. The shRNA transcription units of a plurality of different or same target genes can be connected in series between the two groups of the sequences of the multiple cloning sites of the vector, a plurality of shRNA transcription units thereon can express one target gene for many times or express a plurality of different sites of one target gene or express different target genes after the transcription of cells or animals, so as to specifically inhibit the expressions of two or more target genes. The vector can be easily used in the gene function research and the research and development of the gene therapeutic drugs for diseases.

Description

1. Technical field [0001] The invention relates to an shRNA plasmid expression vector pSilencer-U6 / H1-(shRNA)n based on polygene interference in the field of gene medicine, a construction method and application thereof. 2. Background technology [0002] RNA interference (RNAi) refers to the use of double-stranded RNA (double-stranded RNA, dsRNA) to specifically induce the degradation of mRNA homologous to its sequence, resulting in post-transcriptional gene silencing (PTGS) that specifically inhibits the expression of the corresponding target gene Phenomenon. RNA interference was discovered in 1998 in the genetic analysis of C. elegans. In 1999, it was detected in plants that 21-25nt RNA fragments were necessary for plant RNAi, and they were called small interfering RNA (short or sallinterfering RNA, siRNA). Quickly applied to anti-virus, anti-tumor and gene function research. From 2001 to 2003, RNAi technology was rated as one of the world's top ten scientific achievemen...

AI Technical Summary

Problems solved by technology

The disadvantage of this scheme is that before the construction of multiple shRNA expression vectors, the U6 promoter needs to be isolated and prepared by complex molecular biological means; when the vector is constructed, the isolated U6 promoter needs to be further directionally connected to each of the shRNA expression vectors by enzymatic means. Synthetic "shRNA+U6 termination signal" double-stranded DNA structure, if this operation involves expressing more shRNAs, this directional connection will be more cumbersome

The disadvantage of the patent disclosure scheme of this publication number is that the promoter used is only the H1 promoter, and the commercial initial vector selected as the construction target vector is also limited to pSilencer3.0-H1

Therefore, it is not difficult to conclude that the shRNA / siRNA expression obtained after the expression vector constructed by the patent disclosure scheme of CN1710079A is not as strong as that of the U6 promoter vector, and the resulting target gene expression inhibition effect is not as good as that of the U6 promoter vector. Strong U6 promoter vector

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