کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
2824199 1161629 2012 5 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
A cloning vector for creation of Escherichia colilacZ translational fusions and generation of linear template for chromosomal integration
موضوعات مرتبط
علوم زیستی و بیوفناوری بیوشیمی، ژنتیک و زیست شناسی مولکولی ژنتیک
پیش نمایش صفحه اول مقاله
A cloning vector for creation of Escherichia colilacZ translational fusions and generation of linear template for chromosomal integration
چکیده انگلیسی

A novel cloning vector to aid in the construction of single copy β-galactosidase reporter systems for gene expression studies in lactose metabolizing Escherichia coli strains, including STEC, is described. The plasmid allows construction of translational fusions of cloned gene promoters to a short segment of E. colilacZ. A selectable spectinomycin resistance marker flanked by a short lacI segment is positioned 5′ to the cloning site. PCR amplification using opposing primers complementary to the upstream lacI fragment and the downstream lacZ fragment generates a linear template suitable for integration using pRedET recombination. Integration of linear template derived from the recombinant plasmid into host strains replaces the entire native lacZ promoter and fuses the promoter of interest in-frame with the lacZ gene, thus simultaneously producing a single-copy, chromosomal reporter system and eliminating background lacZ expression. Studies comparing ahpC expression from a chromosomal fusion in the lac open with that on a plasmid in E. coli strain EDL933 are shown.


► A plasmid vector to aid in the construction of β-galactosidase reporter systems.
► A plasmid for gene expression studies in lactose metabolizing Escherichia coli.
► A plasmid for generating lacZ translational fusions for chromosomal integration.

ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Plasmid - Volume 67, Issue 3, May 2012, Pages 259–263
نویسندگان
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