CRISPR/Cpf1 gene editing method applicable to marine crustacean mitochondrion genome

Abstract

The invention discloses a CRISPR/Cpf1 gene editing method applicable to a marine crustacean mitochondrion genome. The method includes the steps: introducing MLS signals positioned by a marine crustacean mitochondrion into a CRISPR/Cpf1 expression plasmid; designing gRNA; micro-injecting fertilized eggs; culturing the fertilized eggs in an in-vitro manner; detecting editing effect to complete editing of the marine crustacean mitochondrion genome. By the method, the marine crustacean mitochondrion genome can be directionally edited, and the method has significant value to a condition that artificial proliferation releasing markers are introduced into the marine crustacean mitochondrion genome.

Description

technical field [0001] The invention belongs to the technical field of molecular biology and relates to a CRISPR / Cpf1 gene editing method suitable for mitochondrial genomes of marine crustaceans. Background technique [0002] Gene editing technology is a technology that modifies specific DNA sequences at the DNA level by means of deletion, insertion, etc. Transcription activator-like effector (TALENs), a qualitative leap has taken place since the establishment of CRISPR / Cas9 technology. On the basis of the CRISPR / Cas9 system, Zhang Feng's research group at the Massachusetts Institute of Technology further improved CRISPR / Cas9 and established the CRISPR / Cpf1 gene editing system. The main difference between it and the CRISPR / Cas9 system is that it uses Cpf1 nuclease instead of Cas9 , but the advantages are more obvious: first, compared with Cas9 nuclease, the Cpf1 enzyme expressed in this system is smaller in size, so it is easier to enter small organelles and tissues such as...

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Problems solved by technology

But apart from this (human), there are no other species, especially research reports on gene editing technology related to the mitochondrial genome of crustaceans