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10.1016/j.stemcr.2015.09.022 http://scihub22266oqcxt.onion/10.1016/j.stemcr.2015.09.022 C4649464!4649464 !26527385     free     free     free Warning: imagejpeg(C:\Inetpub\vhosts\kidney.de\httpdocs\phplern\26527385 .jpg): Failed to open stream: No such file or directory in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 117       Stem+Cell+Reports 2015 ; 5 (5 ): 908-917 Nephropedia Template TP {{tp|p=26527385 |t=2015. Cloning-free CRISPR |pdf=|usr=}}{{26527385 }} gab.com Text Cloning-free CRISPR JO: Stem Cell Reports http://www.kidney.de/pget.php?&tw=1&pmid=26527385 #FOAvip We present self-cloning CRISPR/Cas9 (scCRISPR), a technology that allows for CRISPR/Cas9-mediated genomic mutation and site-specific knockin transgene creation within several hours by circumventing the need to clone a site-specific single-guide RNA (sgRNA) or knockin homology construct for each target locus. We introduce a self-cleaving palindromic sgRNA plasmid and a short double-stranded DNA sequence encoding the desired locus-specific sgRNA into target cells, allowing them to produce a locus-specific sgRNA plasmid through homologous recombination. scCRISPR enables efficient generation of gene knockouts (?88% mutation rate) at approximately one-sixth the cost of plasmid-based sgRNA construction with only 2 hr of preparation for each targeted site. Additionally, we demonstrate efficient site-specific knockin of GFP transgenes without any plasmid cloning or genome-integrated selection cassette in mouse and human embryonic stem cells (2%-4% knockin rate) through PCR-based addition of short homology arms. scCRISPR substantially lowers the bar on mouse and human transgenesis. Twit Text FOAVip Cloning-free CRISPR ????Stem Cell Reports http://www.kidney.de/pget.php?&tw=1&pmid=26527385 #FOAvip Twit Text # Free Paper on # # # # # LINK http://www.kidney.de/pget.php?&tw=1&pmid=26527385 #FOAvip English Wikipedia <ref>{{Cite pmid|26527385 }}</ref> Cloning-free CRISPR #MMPMID26527385 Arbab M ; Srinivasan S ; Hashimoto T ; Geijsen N ; Sherwood RI Stem Cell Reports 2015[Nov]; 5 (5 ): 908-917 PMID26527385 show ga We present self-cloning CRISPR/Cas9 (scCRISPR), a technology that allows for CRISPR/Cas9-mediated genomic mutation and site-specific knockin transgene creation within several hours by circumventing the need to clone a site-specific single-guide RNA (sgRNA) or knockin homology construct for each target locus. We introduce a self-cleaving palindromic sgRNA plasmid and a short double-stranded DNA sequence encoding the desired locus-specific sgRNA into target cells, allowing them to produce a locus-specific sgRNA plasmid through homologous recombination. scCRISPR enables efficient generation of gene knockouts (?88% mutation rate) at approximately one-sixth the cost of plasmid-based sgRNA construction with only 2 hr of preparation for each targeted site. Additionally, we demonstrate efficient site-specific knockin of GFP transgenes without any plasmid cloning or genome-integrated selection cassette in mouse and human embryonic stem cells (2%-4% knockin rate) through PCR-based addition of short homology arms. scCRISPR substantially lowers the bar on mouse and human transgenesis. |*CRISPR-Cas Systems [MESH] |Animals [MESH] |Cells, Cultured [MESH] |Cloning, Molecular [MESH] |Gene Targeting/*methods [MESH] |HEK293 Cells [MESH] |Humans [MESH]Cloning-free CRISPR (epmc).pdf DeepDyve Pubget Overpricing