DEVELOPMENT OF SALT TOLERANT TRANSCENIC MESTA (Hibiscus suhdarjffa L.) PLANT THROUGH AGROBPICTERIUM MEDIATED GENETIC TRANSFORMATION via REGENERATION AND EMBRYOGENESIS

Abstract

Genetic transformation of two gene constructs 11GIy-1 and PsCIPK (that caIn: p131121 of 14KDa plasmid with /i-giucuronidase p35SGLSINT and plain selectable marker gene npttl) confer both salt and drought tolerance to mesta variew t-IS-24 (hibiscus sahi/iviffli) has been undertaken in the Genetic Lingineering Laboratory of Genetic Resources and Seed Division, fliRt. Dhaka-1207. Cotyledons (with attached petioles) and embryos were used as explants br indirect regeneration ob transformed plants and direct transihrmation consecutively. GUS assay was perlormed alter 30 days of cocultivation of infected Cotyledons. GUS positive putative blue color was developed up to 76% ( in case of PsCIPK gene) of cotyledons. But it was observed that 1-15-24 variety was mild transtbrmed (32% (1115 blue color revealed) with PGIy-1 gene construct (cotyledons). GUS positive explants (.Cotyledons) were cultured on shoot regeneration medium (MS medium supplemented with IAA 0.5 mg/I and RAP 2 mg/I) with cefotaxime (500 mg/I). Shoot regeneration percentage oftransfornied plants was 42 to 47 % depending on gene constructs. Transgenic HThLccus .va/ckiri//, plants have also been produced by direct transformation procedure. Embryo axes ol mature seeds were infected by immersion in a suspension of Agmhacicrnun strain culture Following a 24 h cocultivation with .1g/?backhuin strain, embryos were grown on MS medium added with a killer concentration of kananiyein (50 mg/mI) during 4 weeks at room conditions and thereafter translèrred to greenhouse. 76.67% to 83.3410* of the explants excised from putative translbrmed plants were found to be GUS positive depending on gene constructs For salt tolerance testing, \'IS media supplemented with NaCl of different concentrations (50, 75, 100 and 125 nN1) were used. respectively. The results showed the genetic transformation of salt tolerance genes in I 15-24 variety and indicated the possibility of the development oftransgenic niesta (Hibiscus .cahcicui/iz) varieties against salt stresses. From this experiment, the tremendous observation was that Hibiscus scthc/ari/a showed strong response to PsCIPK gene construct but mild response to PGly- I gene construct We also reported here a rapid and efficient method for producing uansgcnic plants rather than regeneration method from H. scthcktriffii embryos, which did not require in ri/lu regeneration.