Identification and molecular cloning of Bm86 gene in Boophilus microplus ticks collected from beef cattle in several regions of Indonesia.

Tick vaccines based on recombinant Bm86 have been used to control the transmission of blood parasites in cattle such as Anaplasma, Babesia, and Theileria. The method is also an alternative to controlling tick infestation a part of chemical pesticide utilization. Bm86 polymorphism is reportedly correlated to the effectiveness of these vaccines. Sequence analysis of Bm86 from Indonesian ticks has not been reported. The aim of this research was to identify, the cloning and expression of the Bm86 gene from several regions of Indonesia (Banten, Sulawesi Selatan, and Yogyakarta) as an initiation to developing a tick vaccine. RNA from female ticks' midgut samples was extracted and followed by cDNA reverse transcription. The cDNA was amplified using Yeerongpilly strain (GenBank Accession number M29321) primer. The Bm86 cDNA amplicon was excised and purified for further sequence analysis. The sequence data were analyzed using MEGA software and BLAST. Bm86 gene was cloned to pET32a and transformed to E. coli BL23. Protein expression was performed by freeze-thawing and sonication methods. Bm86 cDNA from midgut samples were dominantly identical to Bm86 glycoprotein from Thailand (95–98.65% Per Ident). The Bm86 gene was successfully cloned and expressed using the pET32a vector. Bm86 protein was visualized using SDS PAGE that produced a band in 100 kDa. [ABSTRACT FROM AUTHOR]