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10.1038/srep32230 http://scihub22266oqcxt.onion/10.1038/srep32230 C4997560!4997560!27557525     free     free     free Deprecated: Implicit conversion from float 209.6 to int loses precision in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 534 Deprecated: Implicit conversion from float 209.6 to int loses precision in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 534 Deprecated: Implicit conversion from float 209.6 to int loses precision in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 534 Deprecated: Implicit conversion from float 209.6 to int loses precision in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 534 Deprecated: Implicit conversion from float 209.6 to int loses precision in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 534 Deprecated: Implicit conversion from float 209.6 to int loses precision in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 534 Deprecated: Implicit conversion from float 209.6 to int loses precision in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 534       Sci+Rep 2016 ; 6 (ä): ä Nephropedia Template TP {{tp|p=27557525|t=2016. Analysis of gene repair tracts from Cas9/gRNA double-stranded breaks in the human CFTR gene |pdf=|usr=}}{{27557525}} gab.com Text Analysis of gene repair tracts from Cas9/gRNA double-stranded breaks in the human CFTR gene JO: Sci Rep http://www.kidney.de/pget.php?&tw=1&pmid=27557525 #FOAvip To maximise the efficiency of template-dependent gene editing, most studies describe programmable and/or RNA-guided endonucleases that make a double-stranded break at, or close to, the target sequence to be modified. The rationale for this design strategy is that most gene repair tracts will be very short. Here, we describe a CRISPR Cas9/gRNA selection-free strategy which uses deep sequencing to characterise repair tracts from a donor plasmid containing seven nucleotide differences across a 216?bp target region in the human CFTR gene. We found that 90% of the template-dependent repair tracts were >100?bp in length with equal numbers of uni-directional and bi-directional repair tracts. The occurrence of long repair tracts suggests that a single gRNA could be used with variants of the same template to create or correct specific mutations within a 200?bp range, the size of ~80% of human exons. The selection-free strategy used here also allowed detection of non-homologous end joining events in many of the homology-directed repair tracts. This indicates a need to modify the donor, possibly by silent changes in the PAM sequence, to prevent creation of a second double-stranded break in an allele that has already been correctly edited by homology-directed repair. Twit Text FOAVip Analysis of gene repair tracts from Cas9/gRNA double-stranded breaks in the human CFTR gene ????Sci Rep http://www.kidney.de/pget.php?&tw=1&pmid=27557525 #FOAvip Twit Text # Free Paper on # # # # # LINK http://www.kidney.de/pget.php?&tw=1&pmid=27557525 #FOAvip English Wikipedia <ref>{{Cite pmid|27557525}}</ref> Analysis of gene repair tracts from Cas9/gRNA double-stranded breaks in the human CFTR gene #MMPMID27557525 Hollywood JA; Lee CM; Scallan MF; Harrison PT Sci Rep 2016[]; 6 (ä): ä PMID27557525show ga To maximise the efficiency of template-dependent gene editing, most studies describe programmable and/or RNA-guided endonucleases that make a double-stranded break at, or close to, the target sequence to be modified. The rationale for this design strategy is that most gene repair tracts will be very short. Here, we describe a CRISPR Cas9/gRNA selection-free strategy which uses deep sequencing to characterise repair tracts from a donor plasmid containing seven nucleotide differences across a 216?bp target region in the human CFTR gene. We found that 90% of the template-dependent repair tracts were >100?bp in length with equal numbers of uni-directional and bi-directional repair tracts. The occurrence of long repair tracts suggests that a single gRNA could be used with variants of the same template to create or correct specific mutations within a 200?bp range, the size of ~80% of human exons. The selection-free strategy used here also allowed detection of non-homologous end joining events in many of the homology-directed repair tracts. This indicates a need to modify the donor, possibly by silent changes in the PAM sequence, to prevent creation of a second double-stranded break in an allele that has already been correctly edited by homology-directed repair. äAnalysis of gene repair tracts from Cas9/gRNA double-stranded breaks i(epmc).pdf DeepDyve Pubget Overpricing