Expression framework for complex concatenated gRNAs and RNAi for vector expression

Abstract

The invention provides a composite multi-connected gRNA and RNAi expression cassette for vector expression. The composite multi-connected gRNA and RNAi expression cassette can be applied to a eukaryotic expression vector system. The composite multi-connected expression cassette comprises a promoter for promoting transcription, one or more gRNA expression cassette bodies, one or more RNAi expression cassette bodies and a terminator for terminating transcription; the expression cassette has the function of modifying target genes at the DNA level and the mRNA level, and the gRNA / CRISPER gene modifying expression cassette bodies are connected with the RNAi expression cassette bodies through intervening sequences in an alternative series connection mode. The composite multi-connected gRNA and RNAi expression cassette for vector expression solves the problem that in an existing expression structure, one promoter can only be connected with one gRNA and one transcription terminator.

Description

technical field [0001] The present invention relates to an expression framework containing compound and multiple gRNA and RNAi expressed in eukaryotic cells and its construction. Background technique [0002] The CRISPR / Cas system is a new tool for targeted editing of genomic DNA developed in recent years. This system was originally a defense system in bacteria against foreign DNA such as viruses. There are a series of clustered DNA sequences in some bacterial genomes, called "Clustered Regularly Interspaced Short Palindromic Repeats" (CRISPR). The spacers between these repeats were found to be identical to the DNA sequences of many phages capable of invading bacteria. Moreover, after these sequences are transcribed into RNA, they can form a complex with a type of protein called Cas (CRISPR associated) produced by the host bacterium to guide the Cas protein, so this piece of RNA is also called guide RNA (guide RNA, gRNA). When the complex detects that the invading DNA an...

AI Technical Summary

Problems solved by technology

[0009] In order to overcome the shortcomings of the above-mentioned existing single or multiple gRNA expression frameworks and achieve compound, multiple, and convenient gene editing functions, the present invention provides a technical solution, which is a new multi-gRNA and RNAi expression framework, which is characterized by Between a promoter and a terminator in a CRISPR / Cas9 expression vector, there is a "composite, multiple gene editing expression framework" with one or more gRNAs and one or more RNAi

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