کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
6482230 | 1415982 | 2018 | 9 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
Engineering the bacterium Comamonas testosteroni CNB-1: Plasmid curing and genetic manipulation
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کلمات کلیدی
موضوعات مرتبط
مهندسی و علوم پایه
مهندسی شیمی
بیو مهندسی (مهندسی زیستی)
پیش نمایش صفحه اول مقاله
![عکس صفحه اول مقاله: Engineering the bacterium Comamonas testosteroni CNB-1: Plasmid curing and genetic manipulation Engineering the bacterium Comamonas testosteroni CNB-1: Plasmid curing and genetic manipulation](/preview/png/6482230.png)
چکیده انگلیسی
Comamonas species are attractive microbial hosts for bioremediation engineering due to their versatile metabolic capacity and diverse niches inhabited. To establish Comamonas testosteroni CNB-1-a common Comamonas strain-as a programmable cellular chassis, its indigenous 91-kb-long plasmid, pCNB1, must be eliminated; in addition, the organism has to exhibit feasiblity for effective genetic manipulation. Here we present a novel plasmid curing strategy involving rare-cutting homing endonuclease and a selection and counter-selection system. With a self-eliminating helper plasmid, we successfully removed the target plasmid pCNB1, along with the helper plasmid itself, in a single step at an efficiency of 64%. By enhancing the homing endonuclease expression, we were able to further improve the efficiency to almost 100%. The curing of pCNB1 provided a plasmid-free strain for hosting an IncP-type replicon based shuttle vehicle which can be utilized for gene circuit engineering. Additionally, using a PCR-based Cre-loxP system, we demonstrated an increased ability to genetically manipulate the chromosome of C. testosteroni CNB-1. This study provides a rapid and effective solution to eliminate indigenous plasmids, and also facilitates the development of C. testosteroni as a promising chassis for future remediation applications.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Biochemical Engineering Journal - Volume 133, 15 May 2018, Pages 74-82
Journal: Biochemical Engineering Journal - Volume 133, 15 May 2018, Pages 74-82
نویسندگان
Qiang Tang, Ting Lu, Shuang-Jiang Liu,