Two chimeric chromosomes of Streptomyces coelicolor A3(2) generated by single crossover of the wild-type chromosome and linear plasmid scp1

Abstract

Streptomyces coelicolor A3(2) strain 2106 carries a 1.85-Mb linear plasmid, SCP1'-cysD, in addition to a 7.2-Mb linear chromosome. Macrorestriction analysis indicated that both linear DNAs are hybrids of the wild-type chromosome and the linear plasmid SCP1 on each side. Nucleotide sequencing of the fusion junctions revealed no homology between the recombination regions. SCP1'-cysD contains an SCP1 telomere and a chromosomal telomere at each end and therefore does not have terminal inverted repeats. In addition, SCP1'-cysD could not be eliminated from strain 2106 by various mutagenic treatments. Thus, we concluded that both the 7.2-Mb chromosome and SCP1'-cysD are chimeric chromosomes generated by a single crossover of the wild-type chromosome and SCP1. This may be regarded as a model of chromosomal duplication in genome evolution.

FIG. 1.

Comparison of AseI fragments of the linear DNA elements in S. coelicolor A3(2) strains by CHEF assay and Southern hybridization (A) and generation model of SCP1′-cysD and the 7.2-Mb chromosome in strain 2106 (B). (A) (1) CHEF electrophoresis. CHEF electrophoresis was done at 150 V with 90-s pulses for 36 h. Fragment sizes were based on data from genome projects. (2 and 3) Southern hybridization. The 146-kb AseI fragment of SCP1 and the AseI B fragment of the M145 chromosome were used as probes for panels 2 and 3, respectively. (B) Generation model. Black and white bars indicate the SCP1 and chromosomal regions, respectively, with the former enlarged to twice its size. AseI recognition sites, fragment names, and TIRs are also included. The two recombination points are connected by a dashed line.

FIG. 2.

Gene organization (A) and nucleotide sequences (B) around the junctions of SCP1′-cysD and the 7.2-Mb chromosome in strain 2106. (A) pJB9-1 contains the junction of the 7.2-Mb chromosome, and the two arrowheads indicate the primers for PCR amplification of the SCP1′-cysD junction. The names of open reading frames are according to genome projects for S. coelicolor A3(2) and SCP1. SCP1.136* codes for the mutated helicase in SCP1′-cysD. Af, AflII; Ba, BamHI; Cl, ClaI; Sp, SphI; Xh, XhoI. (B) Nucleotide sequences of the corresponding regions of SCP1 and the M145 chromosome. It was revealed that a 55-bp SCP1 DNA and a 15-bp chromosomal DNA were deleted and a T residue was inserted during the recombination process.

FIG. 3.

Southern hybridization analysis of the ends of SCP1′-cysD and the 7.2-Mb chromosome of strain 2106. SCP1, SCP1′-cysD, the M145 chromosome, and the 7.2-Mb chromosome were digested with NcoI, separated by conventional agarose gel electrophoresis, and analyzed by Southern hybridization. pSCP201 (A) and pSUL221 (B) were used as probes for the SCP1 end and the chromosomal end, respectively.