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10.1186/s13036-017-0072-5 http://scihub22266oqcxt.onion/10.1186/s13036-017-0072-5 C5582390!5582390!28878819     free     free     free Deprecated: Implicit conversion from float 219.6 to int loses precision in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 534 Deprecated: Implicit conversion from float 219.6 to int loses precision in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 534 Deprecated: Implicit conversion from float 219.6 to int loses precision in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 534 Deprecated: Implicit conversion from float 219.6 to int loses precision in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 534 Deprecated: Implicit conversion from float 219.6 to int loses precision in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 534 Deprecated: Implicit conversion from float 219.6 to int loses precision in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 534 Deprecated: Implicit conversion from float 219.6 to int loses precision in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 534       J+Biol+Eng 2017 ; 11 (ä): ä Nephropedia Template TP {{tp|p=28878819|t=2017. Construction of an easy-to-use CRISPR-Cas9 system by patching a newly designed EXIT circuit |pdf=|usr=}}{{28878819}} gab.com Text Construction of an easy-to-use CRISPR-Cas9 system by patching a newly designed EXIT circuit JO: J Biol Eng http://www.kidney.de/pget.php?&tw=1&pmid=28878819 #FOAvip Background: Plasmid-borne genetic editing tools, including the widely used CRISPR-Cas9 system, have greatly facilitated bacterial programming to obtain novel functionalities. However, the lack of effective post-editing plasmid elimination methods impedes follow-up genetic manipulation or application. Conventional strategies including exposure to physical and chemical treatments, or exploiting temperature-sensitive replication origins have several drawbacks (e.g., they are limited for efficiency and are time-consuming). Therefore, the demand is apparent for easy and rapid elimination of the tool plasmids from their bacterial hosts after genetic manipulation. Results: To bridge this gap, we designed a novel EXIT circuit with the homing endonuclease, which can be exploited for rapid and efficient elimination of various plasmids with diverse replication origins. As a proof of concept, we validated the EXIT circuit in Escherichia coli by harnessing homing endonuclease I-SceI and its cleavage site. When integrated into multiple plasmids with different origins, the EXIT circuit allowed them to be eliminated from the host cells, simultaneously. By combining the widely used plasmid-borne CRISPR-Cas9 system and the EXIT circuit, we constructed an easy-to-use CRISPR-Cas9 system that eliminated the Cas9- and the single-guide RNA (sgRNA)-encoding plasmids in one-step. Within 3 days, we successfully constructed an atrazine-degrading E. coli strain, thus further demonstrating the advantage of this new CRISPR-Cas9 system for bacterial genome editing. Conclusions: Our novel EXIT circuit, which exploits the homing endonuclease I-SceI, enables plasmid(s) with different replication origins to be eliminated from their host cells rapidly and efficiently. We also developed an easy-to-use CRISPR-Cas9 system with the EXIT circuit, and this new system can be widely applied to bacterial genome editing. Electronic supplementary material: The online version of this article (doi:10.1186/s13036-017-0072-5) contains supplementary material, which is available to authorized users. Twit Text FOAVip Construction of an easy-to-use CRISPR-Cas9 system by patching a newly designed EXIT circuit ????J Biol Eng http://www.kidney.de/pget.php?&tw=1&pmid=28878819 #FOAvip Twit Text # Free Paper on # # # # # LINK http://www.kidney.de/pget.php?&tw=1&pmid=28878819 #FOAvip English Wikipedia <ref>{{Cite pmid|28878819}}</ref> Construction of an easy-to-use CRISPR-Cas9 system by patching a newly designed EXIT circuit #MMPMID28878819 Tang Q; Lou C; Liu SJ J Biol Eng 2017[]; 11 (ä): ä PMID28878819show ga Background: Plasmid-borne genetic editing tools, including the widely used CRISPR-Cas9 system, have greatly facilitated bacterial programming to obtain novel functionalities. However, the lack of effective post-editing plasmid elimination methods impedes follow-up genetic manipulation or application. Conventional strategies including exposure to physical and chemical treatments, or exploiting temperature-sensitive replication origins have several drawbacks (e.g., they are limited for efficiency and are time-consuming). Therefore, the demand is apparent for easy and rapid elimination of the tool plasmids from their bacterial hosts after genetic manipulation. Results: To bridge this gap, we designed a novel EXIT circuit with the homing endonuclease, which can be exploited for rapid and efficient elimination of various plasmids with diverse replication origins. As a proof of concept, we validated the EXIT circuit in Escherichia coli by harnessing homing endonuclease I-SceI and its cleavage site. When integrated into multiple plasmids with different origins, the EXIT circuit allowed them to be eliminated from the host cells, simultaneously. By combining the widely used plasmid-borne CRISPR-Cas9 system and the EXIT circuit, we constructed an easy-to-use CRISPR-Cas9 system that eliminated the Cas9- and the single-guide RNA (sgRNA)-encoding plasmids in one-step. Within 3 days, we successfully constructed an atrazine-degrading E. coli strain, thus further demonstrating the advantage of this new CRISPR-Cas9 system for bacterial genome editing. Conclusions: Our novel EXIT circuit, which exploits the homing endonuclease I-SceI, enables plasmid(s) with different replication origins to be eliminated from their host cells rapidly and efficiently. We also developed an easy-to-use CRISPR-Cas9 system with the EXIT circuit, and this new system can be widely applied to bacterial genome editing. Electronic supplementary material: The online version of this article (doi:10.1186/s13036-017-0072-5) contains supplementary material, which is available to authorized users. äConstruction of an easy-to-use CRISPR-Cas9 system by patching a newly (epmc).pdf DeepDyve Pubget Overpricing