The CRISPR/Cas Nucleases system was used to delete the YY1 binding sites in female mouse ES cells, leaving the CTCF binding site intact. Two target regions were selected: region A positioned upstream of the first YY1 binding site and region B spanning the third site. Positioning the guide RNA target sequence on the site 3 was constrained by the presence of the CTCF site 14 bp downstream. DNA oligonucleotides harboring the sgRNA target sequences were designed using the ZiFiT Targeter program (http://zifit.partners.org/ZiFiT/Disclaimer.aspx ). Oligonucleotide pairs were annealed to generate short double-stranded DNA fragments with overhangs compatible with ligation into the BsmBI-digested plasmid pMLM3636. The Cas9 expression plasmid pJDS246 and the sgRNA expression plasmid pMLM3636 were obtained from Addgene (Addgene #43860 and #47754).
Pjds246
Manufactured by Addgene
PJDS246 is a laboratory product. It is a piece of equipment used in scientific research and experiments. The core function of this product is to perform specific tasks related to the research and analysis conducted in a laboratory setting.
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Lab products found in correlation
2 protocols using pjds246
1
CRISPR-mediated YY1 Binding Site Deletion
2
CRISPR-mediated YY1 Binding Site Deletion
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The CRISPR/Cas Nucleases system was used to delete the YY1 binding sites in female mouse ES cells, leaving the CTCF binding site intact. Two target regions were selected: region A positioned upstream of the first YY1 binding site and region B spanning the third site. Positioning the guide RNA target sequence on the site 3 was constrained by the presence of the CTCF site 14 bp downstream. DNA oligonucleotides harboring the sgRNA target sequences were designed using the ZiFiT Targeter program (http://zifit.partners.org/ZiFiT/Disclaimer.aspx ). Oligonucleotide pairs were annealed to generate short double-stranded DNA fragments with overhangs compatible with ligation into the BsmBI-digested plasmid pMLM3636. The Cas9 expression plasmid pJDS246 and the sgRNA expression plasmid pMLM3636 were obtained from Addgene (Addgene #43860 and #47754).
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