Designing and Cloning Molecular Constructs to Knock Out N-Acetylglucosamine Phosphatidylinositol De-N-Acetylase (GPI12) Gene in Leishmania major (MRHO/IR/75/ER)
GHASEMI NEJAD ALMANI، Pooya نويسنده Leishmaniasis Research Center, Kerman University of Medical Sciences, Kerman, Iran GHASEMI NEJAD ALMANI, Pooya , Sharifi ، Iraj نويسنده Leishmaniasis Research Centre, Kerman University of Medical Sciences, Kerman, Iran , , Kazemi، Bahram نويسنده Kavakeb, P , Babaei Bazkiyaei، Zahra نويسنده , , Bandehpour، Mojgan نويسنده , , Salari Sardoii، Samira نويسنده , , Saedi Dezaki، Ebrahim نويسنده Dept. Of Medical Parasitology, Kerman University Of Medical Science, Kerman ,
Background: Leishmaniasis represents a major public health concern in tropical and sub-tropical countries. At present, there is no efficacious vaccine against the disease and new control methods are needed. One way to access this important goal is to knock out genes of specific macromolecules to evaluate the effect of deletion on the growth, multiplication, pathogenesis and immunity of the parasite. The aim of this study was to design and clone molecular constructs to knock out N-acetylglucosamine phosphatidylinositol de-N-acetylase (GPI12) gene in Leishmania major. Methods: For designing and making molecular constructs, we used pLEXSY-neo2 and pLEXSY-hyg2 vectors. The molecular constructs were cloned in E. coli strain Top10. The molecular constructs were transfected by electroporation into L. major in two stages. Results: The molecular constructs were confirmed by Colony PCR and sequencing. The recombinant strains were isolated by selective antibiotics, after which they were confirmed by PCR, Southern and Western blots. |