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10.1534/genetics.114.169482 http://scihub22266oqcxt.onion/10.1534/genetics.114.169482 C4423380!4423380!25745023     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       Genetics 2015 ; 200 (1): 47-58 Nephropedia Template TP {{tp|p=25745023|t=2015. Avoiding the Ends: Internal Epitope Tagging of Proteins Using Transposon Tn7 |pdf=|usr=}}{{25745023}} gab.com Text Avoiding the Ends: Internal Epitope Tagging of Proteins Using Transposon Tn7 JO: Genetics http://www.kidney.de/pget.php?&tw=1&pmid=25745023 #FOAvip Peptide tags fused to proteins are used in a variety of applications, including as affinity tags for purification, epitope tags for immunodetection, or fluorescent protein tags for visualization. However, the peptide tags can disrupt the target protein function. When function is disrupted by fusing a peptide to either the N or C terminus of the protein of interest, identifying alternative ways to create functional tagged fusion proteins can be difficult. Here, we describe a method to introduce protein tags internal to the coding sequence of a target protein. The method employs in vitro Tn7-transposon mutagenesis of plasmids for random introduction of the tag, followed by subsequent Gateway cloning steps to isolate alleles with mutations in the coding sequence of the target gene. The Tn7-epitope cassette is designed such that essentially all of the transposon is removed through restriction enzyme digestion, leaving only the protein tag at diverse sites internal to the ORF. We describe the use of this system to generate a panel of internally epitope-tagged versions of the Saccharomyces cerevisiae GPI-linked membrane protein Dcw1 and the Candida glabrata transcriptional regulator Sir3. This internal protein tagging system is, in principle, adaptable to tag proteins in any organism for which Gateway-adapted expression vectors exist. Twit Text FOAVip Avoiding the Ends: Internal Epitope Tagging of Proteins Using Transposon Tn7 ????Genetics http://www.kidney.de/pget.php?&tw=1&pmid=25745023 #FOAvip Twit Text # Free Paper on # # # # # LINK http://www.kidney.de/pget.php?&tw=1&pmid=25745023 #FOAvip English Wikipedia <ref>{{Cite pmid|25745023}}</ref> Avoiding the Ends: Internal Epitope Tagging of Proteins Using Transposon Tn7 #MMPMID25745023 Zordan RE; Beliveau BJ; Trow JA; Craig NL; Cormack BP Genetics 2015[May]; 200 (1): 47-58 PMID25745023show ga Peptide tags fused to proteins are used in a variety of applications, including as affinity tags for purification, epitope tags for immunodetection, or fluorescent protein tags for visualization. However, the peptide tags can disrupt the target protein function. When function is disrupted by fusing a peptide to either the N or C terminus of the protein of interest, identifying alternative ways to create functional tagged fusion proteins can be difficult. Here, we describe a method to introduce protein tags internal to the coding sequence of a target protein. The method employs in vitro Tn7-transposon mutagenesis of plasmids for random introduction of the tag, followed by subsequent Gateway cloning steps to isolate alleles with mutations in the coding sequence of the target gene. The Tn7-epitope cassette is designed such that essentially all of the transposon is removed through restriction enzyme digestion, leaving only the protein tag at diverse sites internal to the ORF. We describe the use of this system to generate a panel of internally epitope-tagged versions of the Saccharomyces cerevisiae GPI-linked membrane protein Dcw1 and the Candida glabrata transcriptional regulator Sir3. This internal protein tagging system is, in principle, adaptable to tag proteins in any organism for which Gateway-adapted expression vectors exist. äAvoiding the Ends: Internal Epitope Tagging of Proteins Using Transpos(epmc).pdf DeepDyve Pubget Overpricing