A new approach using multiplex long accurate PCR and yeast artificial chromosomes for bacterial chromosome mapping and sequencing.
An efficient approach for structural studies on bacterial chromosomes is presented. It is based on high-resolution PCR map construction by using a multiplex long accurate PCR (MLA PCR) protocol and a YAC clone carrying the region to be mapped as indicator. The high-resolution PCR map of the bacillus subtilis rrnB-dnaB region is presented as an example. Data are also presented on the use of DNA generated by LA PCR for sequencing; they are relevant to LA PCR induced mutations and justify the application of such mapping for sequencing long stretches of bacterial chromosomes.
Sorokin
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Laboratoire de Génétique Microbienne, Institut National de la Recherche Agronomique, Jouy en Josas, France. sorokine@biotec.jouy.inra.fr
Lapidus
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A
Capuano
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V
Galleron
N
N
Pujic
P
P
Ehrlich
S D
SD
D001412:Bacillus subtilis / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics; D001483:Base Sequence; D002874:Chromosome Mapping / Q000379:methods*; D018244:Chromosomes, Artificial, Yeast / Q000378:metabolism; D002876:Chromosomes, Bacterial; D004265:DNA Helicases; D004269:DNA, Bacterial; D053488:DnaB Helicases; D016521:Electrophoresis, Gel, Pulsed-Field; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods; D017422:Sequence Analysis, DNA / Q000379:methods*; D012407:rRNA Operon
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004265:DNA Helicases; D053488:DnaB Helicases