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Screening method and device for novel CRISPR-Cas system

A new type of candidate region technology, applied in the field of gene editing, can solve problems such as endogenous RNA interference

Active Publication Date: 2018-03-09
SHENZHEN HUADA GENE INST
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] The existing CRISPR-Cas system still has some disadvantages, such as CRISPR-Cas9 requires a special carrier or sacrifices transfection efficiency, and is easily interfered by endogenous RNA in mammalian cells, so it is very important to find a new gene editing system

Method used

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  • Screening method and device for novel CRISPR-Cas system

Examples

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Embodiment 1

[0071] This example is used to prove that the method of the present invention can effectively reduce the number of candidate strains and candidate proteins. In this embodiment, according to figure 1 The method shown was carried out. Specifically, use MetaGeneMark (version 2.8) software to predict the gene sequence and protein sequence of each bacterial strain; use pilecer (version 1.06) software to find the CRISPR region; use interproscan (version 5.16-55.0) software to perform a Annotation; set the first length to 20kb bases and the second length to 500 amino acids, look for proteins with more than 500 amino acids within 20kb bases near the repeat sequence of cas1 or CRISPR region, and extract the proteins that meet the set conditions The protein sequence of the candidate region of the strain; the set conditions include: (a) there is a repeat sequence in the cas1 and CRISPR region, and it does not belong to type I or type III, and the above-mentioned cas1 and the above-menti...

Embodiment 2

[0074] This example verifies the feasibility and efficiency of the method of the present invention. The experimental conditions and parameters of this embodiment are the same as those of Example 1.

[0075] The screening process of the new CRISPR-Cas system is suitable for analyzing the genome data of a single bacterial species, and selecting the strains that may have a new system, which belongs to the second class (Class2) CRISPR-Cas system other than cas9 and cpf1. In order to verify the feasibility and efficiency of the process, the CRISPR-Cas system, type I system, type III system, and CRISPR-cas9 of c2c1, c2c2, and c2c3 non-cas9 and cpf1 (Class 2) were downloaded from the NCBI database. System, CRISPR-cpf1 system, and genome information of strains with both cas9 system and cpf1 system for process verification. A total of 14 strains were validated to verify the feasibility and efficiency of the screening process for the new CRISPR-Cas system.

[0076] 1) Strain informati...

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Abstract

The invention discloses a screening method and device for a novel CRISPR-Cas system. The method comprises the following steps that the predicted gene sequences and protein sequences of strains are provided; CRISPR regions and proteins with cas1 annotation information are acquired; proteins larger than second length near cas1 or repetitive sequences in the range of first length are searched for, and protein sequences in candidate regions of the strains are extracted; a comparison is conducted; annotation result of the highest protein consistency is extracted, highly homologous strains of non 100% comparison ratio with cas9 or cpf1 type are screened out, a secondary structure prediction is conducted, arrangement position information of components of the proteins are acquired, proteins whichare not conformed to cas9 or cpf1 component arrangements are selected to serve as candidate proteins. The method can be used for analyzing single strain genome data so as to select strain proteins which may belong to the novel CRISPR-Cas system.

Description

technical field [0001] The invention relates to the technical field of gene editing, in particular to a method and device for screening novel CRISPR-Cas systems. Background technique [0002] CRISPR (Clustered regularly interspaced short palindromic repeats), known as regularly clustered interspaced short palindromic repeats, is actually a gene editor and a natural immune method in most bacteria and archaea. Through the flanking sequence analysis of the CRISPR cluster, it was found that there was a polymorphic family gene near it, and it worked together with the CRISPR region, so it was named CRISPR-associated gene (CRISPR associated), abbreviated as Cas. Most CRISPR-Cas systems contain cas1 protein, and cas1 is a relatively conserved protein in the Cas family. According to the structure of the effector module, there are mainly two types of CRISPR-Cas systems discovered so far: one class (Class 1) contains multiple Cas proteins and has multiple effector proteins (effectors)...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G06F19/20G06F19/18
CPCG16B20/00G16B25/00
Inventor 李芳杨子翊顾颖李俊桦
Owner SHENZHEN HUADA GENE INST
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