eISSN: 2353-9461
ISSN: 0860-7796
BioTechnologia
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3/2022
vol. 103
 
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abstract:
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Improved transformation of Agrobacterium assisted by silver nanoparticles

Amala Benny
1
,
Swapna Alex
1
,
K.B. Soni
1
,
K.N. Anith
1
,
A.G. Kiran
1
,
M.M. Viji
1

1.
College of Agriculture, Vellayani, Kerala Agricultural University, Kerala, India
BioTechnologia vol. 103(3) ∙ pp. 311–317 ∙ 2022
Online publish date: 2022/09/29
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In transgenic plant development, the low transformation efficiency of Agrobacterium with exogenous DNA is the major constraint, and hence, methods to improve its transformation efficiency are needed. Recently, nanoparticle-mediated gene transfer has evolved as a key transformational tool in genetic transformation. Since silver nanoparticles (AgNPs) can induce pores on the cell membrane, their efficacy in the improvement of conventional calcium chloride freeze-thaw technique of transformation of Agrobacterium was explored in this study. Agrobacterium cells in the exponential growth phase were exposed to different concentrations of AgNPs (0.01, 1, 5, 10, and 20 mg/l), and the half-maximal effective concentration (EC50) was determined via Probit analysis using the SPSS software. Transformation efficiency of AgNPs alone and in combination with calcium chloride was compared with that of the conventional calcium chloride freeze-thaw technique. AgNPs at a concentration of 0.01 mg/l in combination with calcium chloride (20 mM) showed a ten fold increase in the transformation efficiency (3.33 log CFU (colony-forming unit/microgram) of DNA) of Agrobacterium tumefaciens strain EHA 105 with plasmid vector pART27 compared with the conventional technique (2.31 log CFU/μg of DNA). This study indicates that AgNPs of size 100 nm can eliminate the freeze-thaw stage in the conventional (Agrobacterium transformation technique, with a 44% improvement in efficiency. The use of AgNPs (0.01 mg/l) along with 20 mM calcium chloride was found to be an economically viable method to improve the transformation of Agrobacterium with exogenous plasmid DNA.
keywords:

silver nanoparticles, Agrobacterium transformation, calcium chloride, competent cells, freeze thaw, transformation efficiency

 
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