Articles
FACTORS INFLUENCING IN VITRO REGENERATION AND AGROBACTERIUM MEDIATED TRANSFORMATION OF ONION ‘ARKA NIKETAN’
Article number
929_60
Pages
417 – 422
Language
English
Abstract
Onion is an important vegetable crop affected by many diseases and pests.
Developing resistant cultivars can be addressed through gene transfer technology.
However, recalcitrant nature of onion towards in vitro regeneration makes trans-formation a very difficult task.
An optimized protocol for regeneration using embryo explants was developed considering several factors, i.e., the age of the embryogenic callus, the salt content in the basal medium (MS or BDS), gelling agent strength, plant growth regulators (picloram, Kinetin, 2,4-D) in the regeneration medium and aeration of the culture vessels. Agrobacterium mediated transformation was initiated to study the influence of method of inoculation, influence of acetosyringone; and selection agents (kanamycin or geneticin) using Agrobacterium strain LBA 4404 with the binary vector pCambia2301 having nptII gene as selectable marker and GUS gene as the reporter gene under CaMV 35S promoter.
Inoculation of the explants with Agrobacterium under vacuum filtration was suitable only for immature embryos while inoculating with bacterial suspension was suitable for the embryogenic callus.
The effect of acetosyringone was influenced by basal medium and plant growth regulators.
Geneticin was a better selectable agent.
Standardization of transformation was evidenced by optimal transient GUS reaction.
Developing resistant cultivars can be addressed through gene transfer technology.
However, recalcitrant nature of onion towards in vitro regeneration makes trans-formation a very difficult task.
An optimized protocol for regeneration using embryo explants was developed considering several factors, i.e., the age of the embryogenic callus, the salt content in the basal medium (MS or BDS), gelling agent strength, plant growth regulators (picloram, Kinetin, 2,4-D) in the regeneration medium and aeration of the culture vessels. Agrobacterium mediated transformation was initiated to study the influence of method of inoculation, influence of acetosyringone; and selection agents (kanamycin or geneticin) using Agrobacterium strain LBA 4404 with the binary vector pCambia2301 having nptII gene as selectable marker and GUS gene as the reporter gene under CaMV 35S promoter.
Inoculation of the explants with Agrobacterium under vacuum filtration was suitable only for immature embryos while inoculating with bacterial suspension was suitable for the embryogenic callus.
The effect of acetosyringone was influenced by basal medium and plant growth regulators.
Geneticin was a better selectable agent.
Standardization of transformation was evidenced by optimal transient GUS reaction.
Authors
J.B. Mythili, P.R. Rajeev , B.S. Chethana
Keywords
regeneration, transformation, transgenic onion
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