Construction of Gig miR-136 Gene Editing Vector Based on CRISPR/Cas9 Technique
The present paper aims to explore the molecular mechanism of microRNA on the litter size of pig.The expressed vector to edit the miR-136 gene was constructed using the clustered regularly interspaced short palindromic repeat (CRISPR) and associated nuclease Cas9 (CRISPR/Cas9) technique.A pair of single guide RNAs (sgRNA) were designed at the miR-136 gene mature sequence and its downstream location based on the precursor of pig miR-136.Four single DNA oligonucleotides were synthesized with a phosphorylation on each strand of the 5`-end, and annealed into two double-strand oligo fragments: oligo 1 and oligo 2.Both of these oligos were inserted into the BpiI site of plasmid pX462, respectively.After transforming into competent DH5α, the sequences of inserts were confirmed for the two recombinant plasmids as pX462-miR-136-1 and pX462-miR-136-2.Two expression vectors for miR-136 gene editing were constructed, which would be useful for the research of fecundity in pig.
NETL
NSTL
万方数据
维普
