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Vector Construction And Triclosan Selection A The Bla Gene In Puc Which Confersfig

This post categorized under Vector and posted on March 8th, 2020.
PUC19 Vector Cloning: Vector Construction And Triclosan Selection A The Bla Gene In Puc Which Confersfig

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To further validate the utility of FabV-Triclosan selection system we expressed another two proteins namely HIV-1 Nef and HIV-Vif. Vectors pSA-HNef-Bla and pSA-HVif-Bla were modified by replacing bla gene with fabV gene as described in material and methods section. BL21(DE3) were then transformed with pSA-HNef-Bla pSA-HNef-FabV pSA-HVif or Vector construction and triclosan selection.(A) The bla gene in pUC19 which confers ampicillin resistance was replaced with fabI and its promoter region (pFab).The pUC19 multiple cloning site (MCS) is retained however HincII HindIII and PstI are not unique in pFab.The fabI cgraphicette in pFab can be transferred to other pUC-derived plasmids using the AatII and AlwNI restriction sites. To test our hypothesis we exchanged the bla gene in pBR322 vector with fabI. Our hypothesis proved correct when we successfully managed to clone the fabI gene in pBR322 vector in which the fabI was expressed under the control of two promoters i.e. P3 and P1 promoters.

Vector construction and triclosan selection. (A) The bla gene in pUC19 which confers ampicillin resistance was replaced with fabI and its promoter region (pFab). The pUC19 multiple cloning site (MCS) is retained however HincII HindIII and PstI are not unique in pFab. pUC19 is a commonly used plasmid cloning vector in E.coli. The molecule is a small double-stranded circle 2686 base pairs in graphicgth and has a high copy number. pUC19 carries a 54 base-pair multiple cloning site polylinker that contains unique sites for 13 different hexanucleotide-specific restriction endonucleases. pUC19 is one of a series of plasmid cloning vectors created by Joachim Messing and co-workers. The designation pUC is derived from the clgraphicical p prefix (denoting plasmid) and the abbreviation for the University of California where early work on the plasmid series had been conducted.

Vector construction and triclosan selection. (A) The bla gene in pUC19 which confers ampicillin resistance was replaced with fabI and its promoter region (pFab). Use of small amount of non-antibiotic Triclosan as selection agent in growth medium enhanced plasmid stability applicability in various culture media and compatibility with other selection Offering vector creation and gene targeted ES cell lines. Take advantage of our expertise and the services offered by your core facility. Together we design a custom DNA targeting vector for your specific needs.Our scientists handle the complex cloning and verification steps then you work with your core facility to produce your custom mouse model. Overexpression of E. coli endogenous fabI gene and subsequent selection on Triclosan has been proposed as a practical alternative to traditional antibiotic selection systems. Unfortunately overexpression of fabI cannot be used to select medium copy number plasmids typically used for the expression of heterologous proteins in E. coli. Here

PUC19 Vector Cloning Gallery