کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
6494089 | 1418332 | 2018 | 31 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
Iterative genome editing of Escherichia coli for 3-hydroxypropionic acid production
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کلمات کلیدی
موضوعات مرتبط
مهندسی و علوم پایه
مهندسی شیمی
بیو مهندسی (مهندسی زیستی)
پیش نمایش صفحه اول مقاله
چکیده انگلیسی
Synthetic biology requires strategies for the targeted, efficient, and combinatorial engineering of biological sub-systems at the molecular level. Here, we report the use of the iterative CRISPR EnAbled Trackable genome Engineering (iCREATE) method for the rapid construction of combinatorially modified genomes. We coupled this genome engineering strategy with high-throughput phenotypic screening and selections to recursively engineer multiple traits in Escherichia coli for improved production of the platform chemical 3-hydroxypropionic acid (3HP). Specifically, we engineered i) central carbon metabolism, ii) 3HP synthesis, and (iii) 3HP tolerance through design, construction and testing of ~ 162,000 mutations across 115 genes spanning global regulators, transcription factors, and enzymes involved in 3HP synthesis and tolerance. The iCREATE process required ~ 1 month to perform 13 rounds of combinatorial genome modifications with targeted gene knockouts, expression modification by ribosomal binding site (RBS) engineering, and genome-level site-saturation mutagenesis. Specific mutants conferring increased 3HP titer, yield, and productivity were identified and then combined to produce 3HP at a yield and concentration ~ 60-fold higher than the wild-type strain.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Metabolic Engineering - Volume 47, May 2018, Pages 303-313
Journal: Metabolic Engineering - Volume 47, May 2018, Pages 303-313
نویسندگان
Rongming Liu, Liya Liang, Alaksh Choudhury, Marcelo C. Bassalo, Andrew D. Garst, Katia Tarasava, Ryan T. Gill,