Physical Mapping And Phylogetic Analysis Of Babesia Motasi

Babesiosis is a kind of blood protozoonoses caused by a variety of Babesia, which mainly parasitize erythrocytes of cattle, sheep, horses, dogs and other animal and it can lead disease to a variety of vertebrates. It is widespread in tropical, subtropical, temperate regions.Animals infected with Babesia are mainly characterized by fever, emaciation, hemolytic anemia, jaundice and hemoglobinuria. Once infected, it is difficult to eliminate and holds up the development of the world’s cultivation industry. It has made great progress due to recent research on cell invasion, pathopoiesis mechanism, spreading and the diagnosis, prevention and control technology.But the research was less on the molecular mechanisms and gene level.With the emergence of the third generation of high-throughput sequencing technologies and the development of molecular biology, it is possible to conduct the researches at the molecular level especially at genome level.In this study, Babesia motasi lintan and Babesia sp. Xinjiang were selected as the research object, which are widespread in our country. The two Babeisa species were in vitro cultured, and then inoculated the two spleenectomized-sheep. We obtained the integrity of genomic DNA by embedding polypide collected and purified from the merozoite of sheeps red blood in low-melting-point agarose and digesting it with proteinase K.Then we analysised their chromosome size and number by using pulse-field gel electrophoresis and got the phylogenetic tree of the two Babesia by analyzing the single copy genes achieved from third generation of sequencing and the data published in NCBI about the apicomplexan parasites.Finally, we selected four larger Scaffolds according to the genomic sequence data of Babesia motasi Lintan which was sequenced by the third generation of high-throughput sequencing method, and designed 221 pairs of primers with an interval of 50 kb for screening out the bacterial artificial chromosome( BAC) library to get the overlapping relations of the positive clone by DNA fingerprinting.The study showed that the two Babesia species all have four chromosomes and the genome size of B. motasi Lintan is about 11.1M while Babesia sp. Xinjiang is 7.0Mb, which were significantly different and were less than their genomic sequence data.Through the Comparison of the single copy genes, the Babesia sp. Xinjiang have a close relationship with B.bovis, and Babesia motasi Lintan with B.bigemina. Based on the genomic sequence data ofB. Motasi Lintan, 230 positive clones were selected by BAC screening library(about 14000PCR) and fingerprinting verification. We completed the total length of about 11.3Mb of four scaffolds physical map priliminary.The study initially completed the physical map and sequencing data registration verification of Babesia motasi Lin Tan G7, which has laid the solid foundation for the next step of fine physical map and sequence map.