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Optimization of Agrobacterium mediated genetic transformation of cotyledonary node explants of Vigna radiata

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Title Optimization of Agrobacterium mediated genetic transformation of cotyledonary node explants of Vigna radiata
 
Creator Sushil Kumar Yadav
Jyothi Lakshmi Narayana,
Vanaja Maddi
Maheswari Mandapaka
Arun Kumar Shanker
Venkateswarlu Bandi
 
Subject Agrobacterium mediated transformation
Annexin
Double cotyledonary node
Vigna radiata
 
Description Not Available
A reproducible and highly efficient protocol for genetic transformation mediated by Agrobacterium has been
established for greengram (Vigna radiata L. Wilczek). Double cotyledonary node (DCN) explants were inoculated
with Agrobacterium tumefaciens strain LBA 4404 harboring a binary vector pCAMBIA 2301 containing neomycin
phosphotransferase (npt II) gene as selectable marker, β-glucuronidase (GUS) as a reporter (uidA) gene and annexin
1 bj gene. Important parameters like optical density of Agrobacterium culture, culture quantity, infection medium,
infection and co-cultivation time and acetosyringone concentration were standardized to optimize the
transformation frequency. Kanamycin at a concentration of 100 mg/l was used to select transformed cells. Transient
and stable GUS expressions were studied in transformed explants and regenerated putative plants, respectively.
Transformed shoot were produced on regeneration medium containing 100 mg/l kanamycin and 250 mg/l
cefotaxime and rooted on ½ MS medium. Transient and constitutive GUS expression was observed in DCN explants
and different tissues of T0 and T1 plants. Rooted T0 and T1 shoots confirming Polymerase Chain Reaction (PCR)
positive for npt II and annexin 1bj genes were taken to maturity to collect the seeds. Integration of annexin gene
into the greengram genome was confirmed by Southern blotting.
Not Available
 
Date 2021-08-17T09:46:25Z
2021-08-17T09:46:25Z
2012-01-01
 
Type Research Paper
 
Identifier Not Available
Not Available
http://krishi.icar.gov.in/jspui/handle/123456789/57591
 
Language English
 
Relation Not Available;
 
Publisher Springer