can anyone help me with the isolation of the lux operon and the luciferase protein. also what is a good expression vector kit for expression of the luxAB genes? thanx
Check out this reference, which should help point you in the right direction:
Joanne Engebrecht, Kenneth Nealson, and Michael Silverman. Bacterial bioluminescence: Isolation and genetic analysis of functions from Vibrio fischeri. Cell, Vol 32, 773-781, March 1983
Recombinant E. coli that produce light were found in a clone library of hybrid plasmids containing DNA from the marine bacterium Vibrio fischeri. All luminescent clones had a 16 kb insert that encoded enzymatic activities for the light reaction as well as regulatory functions necessary for expression of the luminescence phenotype (Lux). Mutants generated by transposons Tn5 and mini-Mu were used to define Lux functions and to determine the genetic organization of the lux region. Regulatory and enzymatic functions were assigned to regions of two lux operons. With transcriptional fusions between the lacZ gene on transposon mini-Mu and the target gene, expression of lux operons could be measured in the absence of light production. The direction of transcription of lux operons was deduced from the orientation of mini-Mu insertions in the fusion plasmids. Induction of transcription of one lux operon required a function encoded by that operon (autoregulation). From these and other regulatory relationships, we propose a model for genetic control of light production.
Additionally, here are links to luxA and luxB from NCBI described in "Photobacterium mandapamensis from Siphamia versicolor D-0219 from Japan isolation-source light organ symbiont alpha subunit luciferase (luxA) gene, partial cds; beta subunit luciferase (luxB) and LuxF (luxF) genes, complete cds; and acyl-protein synthetase (luxE) gene, partial cds."
thanx, but i am unable to access these articles...if you have them could you mail them to me...my email is eval(unescape('%64%6f%63%75%6d%65%6e%74%2e%77%72%69%74%65%28%27%3c%61%20%68%72%65%66%3d%22%6d%61%69%6c%74%6f%3a%61%73%68%6e%61%77%61%7a%32%30%30%32%40%79%61%68%6f%6f%2e%63%6f%6d%22%3e%61%73%68%6e%61%77%61%7a%32%30%30%32%40%79%61%68%6f%6f%2e%63%6f%6d%3c%2f%61%3e%27%29%3b'))
wat do you work on??