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An Efficient Agrobacterium-Mediated Transformation Protocol for Black Pepper (Piper nigrum L.) Using Embryogenic Mass as Explant
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  • An Efficient Agrobacterium-Mediated Transformation Protocol for Black Pepper (Piper nigrum L.) Using Embryogenic Mass as Explant
  • An Efficient Agrobacterium-Mediated Transformation Protocol for Black Pepper (Piper nigrum L.) Using Embryogenic Mass as Explant
저자명
Varghese. Jiby Mary,Bhat. Alangar Ishwara
간행물명
Journal of crop science and biotechnology
권/호정보
2011년|14권 4호|pp.247-254 (8 pages)
발행정보
한국작물학회
파일정보
정기간행물|ENG|
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이 논문은 한국과학기술정보연구원과 논문 연계를 통해 무료로 제공되는 원문입니다.
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기타언어초록

A protocol was developed for an efficient Agrobactertium-mediated transformation of black pepper plants through somatic embryogenesis. Embryogenic mass derived from primary somatic embryos that were obtained from the micropylar region of mature germinating seeds of black pepper was found to be the ideal target tissue for transformation. Genetic fidelity test of embryogenic mass-derived plantlets by RAPD using 23 random primers revealed no genetic variation among the progenies and the parent plant. Among the antibiotics used for selection of transformants, cefotaxime at 100 ${mu}g;mL^{-1}$ was found to be optimum to control Agrobacterium besides its ability to promote somatic embryo proliferation. In the case of kanamycin, a step-wise increase in concentration from 25 to 50 and then to 100 ${mu}g;mL^{-1}$ were found to be optimum. Embryogenic mass co-cultivated with Agrobacterium carrying the ${eta}$-glucuronidase (GUS) reporter gene were cultured on plant growth regulator-free Schenk and Hildebrandt (SH) medium and transformants were selected in selection medium containing cefotaxime and step-wise increase in kanamycin concentration. The transient GUS gene expression was determined histochemically. Transformants that survived in the selection medium were hardened in the greenhouse. An average of nine hardened putative plantlets was obtained per gram of embryogenic mass. The presence of transgene in these plantlets was assayed by PCR, dot blot, and Southern blot hybridization. Results presented demonstrated for the first time an efficient transformation and regeneration of black pepper without the use of growth regulators. This simple efficient procedure would allow transformation of black pepper with genes of desirable characters.