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10.1038/ncomms10431 http://scihub22266oqcxt.onion/10.1038/ncomms10431 C4736110!4736110!26786405     free     free     free Deprecated: Implicit conversion from float 217.6 to int loses precision in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 534 Deprecated: Implicit conversion from float 217.6 to int loses precision in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 534 Deprecated: Implicit conversion from float 217.6 to int loses precision in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 534 Deprecated: Implicit conversion from float 217.6 to int loses precision in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 534       Nat+Commun 2016 ; 7 (ä): ä Nephropedia Template TP {{tp|p=26786405|t=2016. ssODN-mediated knock-in with CRISPR-Cas for large genomic regions in zygotes |pdf=|usr=}}{{26786405}} gab.com Text ssODN-mediated knock-in with CRISPR-Cas for large genomic regions in zygotes JO: Nat Commun http://www.kidney.de/pget.php?&tw=1&pmid=26786405 #FOAvip The CRISPR-Cas system is a powerful tool for generating genetically modified animals; however, targeted knock-in (KI) via homologous recombination remains difficult in zygotes. Here we show efficient gene KI in rats by combining CRISPR-Cas with single-stranded oligodeoxynucleotides (ssODNs). First, a 1-kb ssODN co-injected with guide RNA (gRNA) and Cas9 messenger RNA produce GFP-KI at the rat Thy1 locus. Then, two gRNAs with two 80-bp ssODNs direct efficient integration of a 5.5-kb CAG-GFP vector into the Rosa26 locus via ssODN-mediated end joining. This protocol also achieves KI of a 200-kb BAC containing the human SIRPA locus, concomitantly knocking out the rat Sirpa gene. Finally, three gRNAs and two ssODNs replace 58-kb of the rat Cyp2d cluster with a 6.2-kb human CYP2D6 gene. These ssODN-mediated KI protocols can be applied to any target site with any donor vector without the need to construct homology arms, thus simplifying genome engineering in living organisms. Twit Text FOAVip ssODN-mediated knock-in with CRISPR-Cas for large genomic regions in zygotes ????Nat Commun http://www.kidney.de/pget.php?&tw=1&pmid=26786405 #FOAvip Twit Text # Free Paper on # # # # # LINK http://www.kidney.de/pget.php?&tw=1&pmid=26786405 #FOAvip English Wikipedia <ref>{{Cite pmid|26786405}}</ref> ssODN-mediated knock-in with CRISPR-Cas for large genomic regions in zygotes #MMPMID26786405 Yoshimi K; Kunihiro Y; Kaneko T; Nagahora H; Voigt B; Mashimo T Nat Commun 2016[]; 7 (ä): ä PMID26786405show ga The CRISPR-Cas system is a powerful tool for generating genetically modified animals; however, targeted knock-in (KI) via homologous recombination remains difficult in zygotes. Here we show efficient gene KI in rats by combining CRISPR-Cas with single-stranded oligodeoxynucleotides (ssODNs). First, a 1-kb ssODN co-injected with guide RNA (gRNA) and Cas9 messenger RNA produce GFP-KI at the rat Thy1 locus. Then, two gRNAs with two 80-bp ssODNs direct efficient integration of a 5.5-kb CAG-GFP vector into the Rosa26 locus via ssODN-mediated end joining. This protocol also achieves KI of a 200-kb BAC containing the human SIRPA locus, concomitantly knocking out the rat Sirpa gene. Finally, three gRNAs and two ssODNs replace 58-kb of the rat Cyp2d cluster with a 6.2-kb human CYP2D6 gene. These ssODN-mediated KI protocols can be applied to any target site with any donor vector without the need to construct homology arms, thus simplifying genome engineering in living organisms. ässODN-mediated knock-in with CRISPR-Cas for large genomic regions in z(epmc).pdf DeepDyve Pubget Overpricing