Genetic transformation of chickpea (Cicer arietinum L.) with insecticidal crystal protein gene using particle gun bombardment.
Here, we report the establishment of an efficient particle gun bombardment mediated genetic transformation in chickpea (Cicer arietinum L.) using cryIAc gene of Bacillus thuringiensis. Explants were bombarded with recombinant plasmids engineered for the expression of cryIAc transgene in plants and stable transformants regenerated in presence of benzyladenine, kinetin and kanamycin. Transformation frequency showed dependence on explant type, cultivars, plasmids, helium pressure and microcarrier type used. Integration of transgenes was demonstrated using polymerase chain reaction and Southern blot hybridization approaches in T (0) plants. The expression of CryIA(c) delta-endotoxin and GUS enzyme was ascertained by enzyme linked immunosorbent assay and histochemical assays, respectively. These transgenic plants (T (0)) showed more protection and high mortality for Heliothis armigera and Spodoptera litura larvae as compared to control plants. The results of the present study indicate that highest transformation frequency (18%) could be achieved by use of gold as a microcarrier in combination with helium pressure of 900 psi. Among the other factors tested, plasmid pHS 102 was the most efficient plasmid, while epicotyl explant was the best explant source for particle gun bombardment. Among the different cultivars of chickpea tested, cultivar ICCC37 and PG-12 produced higher frequency of transformation frequency compared to others.
Indurker
Shivani
S
Nuclear Agriculture and Biotechnology Division, Bhabha Atomic Research Centre, Mumbai, Maharashtra, India.
Misra
Hari S
HS
Eapen
Susan
S
D000818:Animals; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000235:genetics*; D001427:Bacterial Toxins / Q000235:genetics*; D001483:Base Sequence; D019470:Biolistics; D029863:Cicer / Q000235:genetics*; D017931:DNA Primers; D004731:Endotoxins / Q000235:genetics*; D004797:Enzyme-Linked Immunosorbent Assay; D005247:Feeding Behavior; D006460:Hemolysin Proteins / Q000235:genetics*; D009036:Moths / Q000502:physiology; D030821:Plants, Genetically Modified; D016133:Polymerase Chain Reaction; D014170:Transformation, Genetic
D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D001427:Bacterial Toxins; D017931:DNA Primers; D004731:Endotoxins; D006460:Hemolysin Proteins; C030956:insecticidal crystal protein, Bacillus Thuringiensis