C. jejuni ORF library
Campylobacter jejuni is the most commonly recognised
cause of bacterial gastroenteritis in humans. Initial difficulties
in culture requirements, and attempts at genetic modification held
back the progress in understanding this organism compared to other
enteric pathogens. However, the sequencing of 1.64 million
nucleotides in the genome at the Wellcome Trust Sanger Institute¹
has heralded a new era in C. jejuni research. Using this
sequence information, along with 31 predicted ORF's by TIGR, Jodi
Parrish (Russell Finley's lab - Wayne State University School of
Medicine) has created a C. jejuni ORF library². The
proteome-scale expression clone set comprises of >1,600 ORF's.
The clones were generated by efficiently utilizing the native E.
coli recombination system. Recombination sites have been confirmed
by sequencing, and most of the clones have been 5' sequence
verified. The ORFs have a range from 45 - 4020bp in length. After
expression, affinity purification can be performed on the proteins
using either GST or His-tags. The clone set has many applications,
including protein interactions, activity, and structural studies.
Furthermore, the clones can be utilised for gene expression
patterns by micro-array analysis.
This is the diagrammatic representation of the expression and
cloning regions of pTLJ03 ( vector map, sequence). The process
of endogenous recombination in E. coli facilitates the
insertion of the ORF in the correct 5' to 3' direction. The
antibiotic resistance gene, encoded on the plasmid, allows for the
selection of the new construct.
Format
Source BioScience LifeSciences offers the complete clone set of
17 96-well microtitre plates or individual clones, for research
purposes only. There is also a C. jejuni
finder tool available which allows you to find the location of the
clone in the 96-well plates. We can also make subsets of your
chosen clones from the collection. Please contact
us to discuss your requirements.
References.
¹Parkhill J, Wren BW, Mungall K, Ketley JM, Churcher C, Basham
D, Chillingworth T, Davies RM, Feltwell T, Holroyd S, Jagels K,
Karlyshev AV, Moule S, Pallen MJ, Penn CW, Quail MA, Rajandream MA,
Rutherford KM, van Vliet AH, Whitehead S, Barrell BG. The genome
sequence of the food-borne pathogen Campylobacter jejuni
reveals hypervariable sequences. Nature. 2000 Feb 10; 403 (6770):
665-8.
²Parrish J, Limjindaporn T, Hines JA, Liu J, Liu G, and Finley RL,
Jr. High-throughput cloning of Campylobacter jejuni ORFs by in vivo
recombination in Escherichia coli, J. Proteome Research, in press
(2004).