title: A novel prokaryotic vector for identification and selection of recombinants: direct use of the vector for expression studies in E. coli.
abstract: BACKGROUND
  The selection of bacterial recombinants that harbour a desired insert, has been a key factor in molecular cloning and a series of screening procedures need to be performed for selection of clones carrying the genes of interest. The conventional cloning techniques are reported to have problems such as screening high number of colonies, generation of false positives, setting up of control ligation mix with vector alone etc.
  
  
  RESULTS
  We describe the development of a novel dual cloning/expression vector, which enables to screen the recombinants directly and expression of the gene of interest. The vector contains Green fluorescence protein (GFP) as the reporter gene and is constructed in such a way that the E. coli cells upon transformation with this vector does not show any fluorescence, but readily fluoresce upon insertion of a foreign gene of interest. The same construct could be easily used for screening of the clones and expression studies by mere switching to specific hosts.
  
  
  CONCLUSIONS
  This is the first vector reported that takes the property of colour or fluorescence to be achieved only upon cloning while all the other vectors available commercially show loss of colour or loss of fluorescence upon cloning. As the fluorescence of GFP depends on the solubility of the protein, the intensity of the fluorescence would also indicate the extent of solubility of the expressed target protein.
authors.affiliation: Lupin Limited, Biotechnology R & D, Gat #1156, Ghotawade Village, Mulshi Taluka, Pune-411042, India. srirampadmanabhan@lupinpharma.com
authors.forename: Sampali
authors.initials: S
authors.lastname: Banerjee
authors.forename: Jitendra
authors.initials: J
authors.lastname: Kumar
authors.forename: Anjali
authors.initials: A
authors.lastname: Apte-Deshpande
authors.forename: Sriram
authors.initials: S
authors.lastname: Padmanabhan
mesh_terms: D003001:Cloning, Molecular / Q000379:methods*; D004926:Escherichia coli / Q000235:genetics*; D017930:Genes, Reporter / Q000235:genetics; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000235:genetics; D015336:Molecular Probe Techniques; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000302:isolation & purification; D012995:Solubility
chemical_list: D011994:Recombinant Proteins; D049452:Green Fluorescent Proteins
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