[Construction of nsdAmgh gene disruption mutant in Strempomyces roseoflavus Men-myco-93-63].

Insertional mutagenesis is a widely used method to determine the function(s) of a gene. To study the function(s) of the gene nsdAmgh in Streptomyces roseoflavus, a homologous recombination vector pSRNA2500 was structured in this paper. The recombination donor vector was then transformed into Strempomyces roseoflavus strain Men-myco-93-63 by conjugative transfer. The transformants were subjected to selection under the pressure of high temperature and appropriate antibiotics. As a result, several disrupted mutants of nsdAmgh gene, with a phenotype of Am(s)Km(r), were isolated and verified using PCR and Dot-blotting and Southern blotting hybridization methods. Functional analysis showed that the disrupted mutants of nsdAmgh had a two-fold higher inhibition against Verticillium dahlia Kleb than that of the wild strain Men-myco-93-63, which all will provide a new study route for future research about positive and negative regulator in Men-myco-93-63.

A plant-transformation-competent BIBAC/BAC-based map of rice for functional analysis and genetic engineering of its genomic sequence.

Sequencing of the rice genome has provided a platform for functional genomics research of rice and other cereal species. However, multiple approaches are needed to determine the functions of its genes and sequences and to use the genome sequencing results for genetic improvement of cereal crops. Here, we report a plant-transformation-competent, binary bacterial artificial chromosome (BIBAC) and bacterial artificial chromosome (BAC) based map of rice to facilitate these studies. The map was constructed from 20 835 BIBAC and BAC clones, and consisted of 579 overlapping BIBAC/BAC contigs. To facilitate functional analysis of chromosome 8 genomic sequence and cloning of the genes and QTLs mapped to the chromosome, we anchored the chromosomal contigs to the existing rice genetic maps. The chromosomal map consists of 11 contigs, 59 genetic markers, and 36 sequence tagged sites, spanning a total of ca. 38 Mb in physical length. Comparative analysis between the genetic and physical maps of chromosome 8 showed that there are 3 "hot" and 2 "cold" spots of genetic recombination along the chromosomal arms in addition to the "cold spot" in the centromeric region, suggesting that the sequence component contents of a chromosome may affect its local genetic recombination frequencies. Because of its plant transformability, the BIBAC/BAC map could provide a platform for functional analysis of the rice genome sequence and effective use of the sequencing results for gene and QTL cloning and molecular breeding.