Analysis Of Insertion Sequence Complement In Bacillus Thuringiensis YBT-1520

Insertion sequences (ISs) are small transposable DNA fragments that can insert into one or multiple loci of genome.It is well recognized that IS elements sometimes inactivate genes upon insertion or activate and/or enhance the expression of nearby genes. Meanwhile, ISs have played an important role in genome reshuffling and evolution by facilitating horizontal gene transfer and mediating homologous recombination between multiple copies present in a given genome.Sequencing of YBT-1520 genome revealed 134 transposase coding genes (at least 68 intact genes) on the chromosome which is one of the most notable features of YBT-1520. In this work,fourteen kinds of intact ISs including seven new-named elements were identified in YBT-1520.A detailed characterization of these ISs distributed into 6 families is presented. Moreover, a comparative analysis of their counterparts in 18 finished Bacillus cereus group genomes as well as in different B. thuringiensis strains has been done in order to understand the distribution and evolution of these IS elements.The IS elements of YBT-1520 were subjected to a careful analysis of IS copies, location and insertion into function genes.Furthermore, these ISs were compared to the elements of the other B. cereus group genomes, in an attempt to demonstrate the distribution and evolution of IS elements in these closely related genomes. YBT-1520 chromosome contains a larger number of transposase coding genes than that of other B. cereus group genomes mainly due to the multi-copies of IS231C, IS232A and ISBth166. Although most of the IS elements randomly distributed throughout the genome were inserted in non-coding regions, at least 9 CDSs were disrupted by the insertion of IS elements (mostly by IS231C). The only copy of siderophore biosynthesis protein coding gene, which is a growth-determining factor under iron-limited conditions in the host insect gut, was inserted by IS232A.To further assess the involvement of IS elements in promoting genome reorganization between two B. thuringiensis kurstaki strains (YBT-1520 and B. thuringiensis HD-1),hybridization experiments were performed using fragments of three most abundant IS elements (IS231C, IS232A and ISBth166) of different families as probes. The result showed that the most multitudinous IS231C had minor different hybridization profiles between these two isolates, which indicated that it did not cause large-scale rearrangements.The distribution of ISBth166 among B. thuringiensis serovars was further examined by Southern blot analysis which suggesting that ISBth166 is widely distributed among kurstaki strains.Genome survey of B. thuringiensis CT-43 revealed at least 40 intact IS elements. Multi-copies of IS231A, in stead of IS231C, were found in CT-43 genome. Meanwhile,4 copies of ISs show 87% similarity to ISBth166.All these results showed species specificity of ISs in B. thuringiensis and was in concert with the conclusion we've drawn before.To data, eighteen B. cereus group genomes have been completely sequenced and published in Genbank. A careful analysis and re-annotation were performed to the IS complement in B. cereus group. The extended survey of all IS elements among the B. cereus group genomes showed some correlation between MLST phylogenetic relatedness and distribution of IS families.IS3 family transposases of subgroup I members were clustered together. The different existence of IS4 and IS110 family elements were also found between subgroup I and the other two groups. This indicated that the IS elements of subgroup I B. anthracis-related genomes may have their distinct phylogenetic lineage, which may affected by their different ecological niche. The disequilibrium in distribution and copy numbers of ISs among B. cereus group genomes is probably one of the major forces of genome evolution.Genome analyses of cytochrome P450 complement of Nocardia farcinica genome were further performed using the same method. Comparative analyses of these cytochrome P450 proteins with their courterparts in related Actinomycetes showed different function relationship among them. The unique features of Nocardia farcinica genome were probably due to the abundant and various cytochrome P450 members.