Detecting the presence of GRP-78 gene in syrian strain of leishmania tropica genome and sequencing it

Abstract

Background: Leishmania tropica is the main cause of cutaneous leishmaniasis which represents a common issue in Syria. After the start of the current war, cutaneous leishmaniasis has become yet more widespread, so an effective vaccine is needed to prevent a largescale spread of leishmania in the country. DNA vaccination is a promising approach for developing such needed vaccine against leishmaniasis. Glucose-regulated protein 78 plays important roles in Leishmania parasite. The gene that codes this protein had been used as DNA vaccine against infections by different species of Leishmania, so this gene represents a good candidate for DNA vaccine designing. But the existence of glucose-regulated protein 78 gene has not yet been proven in the genome of Leishmania tropica. So, this study aims to detect the existence and the expression of glucoseregulated protein 78 gene in Syrian strain of Leishmania tropica promastigotes. Materials & methods: The DNA and RNA were isolated from Syrian strain of Leishmania tropica parasites, the cDNA was synthesized, necessary primers for genes’ amplification were designed manually, the PCR conditions were optimized. Full-length double-stranded sequence analysis was performed on PCR products from cDNA. Results: Extracted genomic DNA and total RNA showed high degree of purity and integrity. GRP78 PCR products showed by gel electrophoresis, on the level of DNA and cDNA, only one band belongs to the gene, and their sizes were approximately 1977 bp. The sequence of the cDNA-GRP78 gene was determined and published in GenBank, and the precise size of the gene was 1977 bp. Expression was also demonstrated at the level of cDNA. Conclusion: Glucose-regulated protein 78 gene is a part of Syrian strain of L. tropica, the expression of this gene has been demonstrated. The sequence of the cDNA of glucose-regulated protein 78 gene has defined and submitted to the Genbank with accession number NO. MN458469