An Intergeneric Conjugation System And The Preliminary Trial For The Construction Of The Chromosome Physical Map Of S. Nanchangensis NS3226

In this study, the suitable parameters for the introduction of plasmids pHZ1358 and pSET152 into S. nanchangensis NS3226 from E. coli were tested for developing an conjugation system for S. nanchangensis NS3226.A dnd gene cluster, which encodes an unknown modification system for 5 Hvidans 1326 and renders its DNA susceptible to site-specific double-strand DNA cleavage during electrophoresis was conjugated from E. coli into S. nanchangensis NS3226. The total DNA of exoconjugants acquired DNA degradation phenotype during electrophoresis, which suggested that the dnd gene cluster was heterologously expressed. The pHZ1358 was used sucessfully as a vector for gene replacement to identify the biosynthetic pathway genes for nanchangmycin in S. nanchangensis NS3226. However, the gene replacement in another potential PKS cluster has not succeeded till now.The overall structure of the chromosome of S. nanchangensis NS3226was shown to be linear DNA molecule with covalently bound terminal proteins. The chromosome telomeres of this strain were seemingly to lie on the two largest chromosomal AseI fragments, but the conclusion needs to be refined.The work on physical mapping of the chromosome of S. nanchangensis NS3226 was initiated. Nearly a full set of chromosomal AseI-BamHI fragments of S. nanchangensis NS3226 were cloned and used as probe to hybridized against its genomic library. Thirty four AseI linking cosmids were observed from 162 hybridizing cosmids and 20 of them showed no obvious overlapping each other by BamHI digestion, suggesting distinct identifications.