Method for realizing animal sex control by editing X chromosome multi-copy genes
An X chromosome and multi-copy technology, applied in the field of genetic engineering, can solve the problems of gender control effect to be further improved and high off-target rate, achieve good sex control effect, low off-target rate, and increase birth rate
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[0039] 1. Selection of mouse multi-copy gene target sites and design of corresponding sgRNA
[0040] The X staining of the mouse was deeply sequenced, and the repeated sequences with more than 50 copies were selected for use. Then, the CRISPR / Cas9 target sequence online design software was used to analyze the candidate sequence, and compared with the whole mouse genome sequence, the X-B and X-D sites with the lowest off-target efficiency were selected, and the corresponding sgRNA X-B and sgRNA X-D sequences were designed. The information is shown in Table 1-2 below.
[0041] Table 1 Multi-copy target loci on mouse X chromosome
[0042] target site name Sequence (5'-3' underline marked as PAM region) copy number X-B GCTTGGTTAGGGTGAGTGCT TGG (SEQ ID NO:1)
72 X-D TAAGTGCTGTGTGCTGCTAC TGG (SEQ ID NO:2)
99
[0043] Table 2 sgRNA sequence
[0044] sgRNA name target site Sequence (5'-3') sgRNA X-B X-B GCTTGGTTAGGGT...
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