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A method for designing a CRISPR-induced RNA library

A design method and library technology, applied in genomics, instrumentation, sequence analysis, etc., can solve the problems of impracticality and high computational cost

Active Publication Date: 2021-04-09
CENT SOUTH UNIV
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Problems solved by technology

However, the calculation cost of GuideScan is relatively high, especially when calculating the off-target sequence of the induced RNA (the number of mismatches with the induced RNA is not less than the parameter M and not greater than the parameter Q, Q>M), the calculation cost of GuideScan is very high
Therefore, it is impractical to apply GuideScan to large genomes

Method used

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Embodiment Construction

[0038] The present invention will be further described below in conjunction with examples.

[0039] This embodiment provides a method for designing a CRISPR-induced RNA library, which specifically includes the following steps:

[0040] Step 1: Determine the targetable genomic space in parallel.

[0041] In order to generate the induced RNA library desired by the user, the method also allows the user to input the reference genome FASTA file, the length of the induced RNA, the standard PAM (protospacer adjacent motif), the PAM position relative to the induced RNA target sequence, non- Standard PAM, and Hamming distances M and Q. Such as figure 1 As shown in (b, c), based on the parameters given by the user, the algorithm scans the kmer prefixed or suffixed by standard PAM and non-standard PAM in the reference genome, as well as related information, such as its corresponding coordinates and directions (used to record kmer in position and encoding direction in a reference seque...

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Abstract

The invention discloses a method for designing a CRISPR-induced RNA library, comprising the following steps: step 1, generating a kmer set according to a reference genome; step 2, dividing kmer into kmer1 and kmer2, and dividing corresponding kmer1 and kmer2 into one category ; Then the kmer2 of the same category is built into the same retrieval tree, and the key sequence of each retrieval tree is kmer1 corresponding to kmer2; Step 3, obtaining inducible RNA and its off-target sequence in parallel, in this step, comparing a kmer and When the key sequence of the search tree is connected with the kmer of kme2, first compare the kmer1 of the kmer with the key sequence of the search tree to see if the set conditions are met, then continue to compare the kmer2 of the kmer with the key sequence of the search tree kmer2. The invention improves computing efficiency.

Description

technical field [0001] The invention belongs to the field of functional genomics, in particular to a method for designing a CRISPR-induced RNA library. Background technique [0002] Among the current generation of genome editing technologies, CRISPR system (Clustered regularly interspaced short palindromic repeats clustered regularly interspaced short palindromic repeats) and Cas9 nuclease (RNA guide nuclease 9 associated with CRISPR) are developing fastest , which can be easily targeted to almost any genomic location, which is a big step forward for the innovative development of genetic engineering. The Cas9 protein from the CRISPR system localizes the complex of inducible RNA and protein to the DNA target sequence by utilizing the characteristics of base pairing between the inducible RNA (guide RNA, gRNA, also known as guide RNA) and the DNA target sequence. Binding to a protospacer-adjacent motif (PAM) downstream of the target site helps guide Cas9 to cut double-strand D...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G16B20/00G16B30/10
CPCG16B20/00G16B30/10
Inventor 王建新李涛王劭恺严承李敏
Owner CENT SOUTH UNIV
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