Global Advanced Research Journal of Agricultural Science (GARJAS) ISSN: 2315-5094
April 2017 Vol. 6(4): pp. 097-107
Copyright © 2017 Global Advanced Research Journals

 

Full Length Research Paper

Factors Affecting Agrobacterium-mediated Transformation and Callus  Induction of Sudan’s Cotton Genotypes

Iman K.A. Abdel Gadir 1,2, Marmar A. El Siddig3*, Hayfa H.A. Ibrahim3 and Adil A. El Hussein3

 

1Faculty of Science, Jazan University, Saudi Arabia;

2Faculty of Science and Technology, Shendi University, Sudan;

3Faculty of Science, University of Khartoum, Sudan

Email: marmarelsiddig@hotmail.com

Accepted 10 April, 2017

 

Abstract

The objective of this study was to optimize conditions affecting transformation of insect resistance gene (cry 1Ac) into locally grown cotton cultivars (Hamid, Barakat- 90) using a binary Agrobacterium tumefaciens vector. The highest percentages (99.33 and 85.67%) of callus induction from hypocotyls of Hamid and Barakat- 90, respectively were observed in MS medium supplemented with 2 mg/L 2-isopentynyl adenine (2iP) + 2 mg/ L IAA. Transformation/callus induction protocol for cotton cultivars were developed using A. tumefaciens EHA105 strain harboring the pCAMBIA 2301 vector which contains a cry 1Ac gene coding for insecticide resistance under the transcriptional control of cauliflower mosaic virus 35S promoter. Results showed that 16.1—22.3% of the co-cultivated explants were Hygromycin resistant and were considered as putative transformants. Transformation was further confirmed by GUS histochemical assay and PCR amplification of the 35S promoter, Nos terminator, Hygromycin phosphotransferase (hpt) and Kanamycin resistance (nptII) genes which were found on the inserted construct. GUS expression was observed in transformed cotton calli but never in the non-transformants (control). Also, transformed calli gave the target PCR amplicons for all of the examined genes while the untransformed tissues didn’t. Transformation percentages of 16.7 for Hamid and 11.9 for Barakat- 90 were reported. 

Keywords: Transformation, callus, cotton, Agrobacterium tumefaciens.


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