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African Journal of Biotechnology

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Factor affecting Agrobacterium-mediated transformation of rice chitinase gene in Solanum tuberosum L.

Muhammad Zulfiqar Ahmad, Iqbal Hussain, Aish Muhammad, Shaukat Ali, Ghulam Muhammad Ali, Sohaib Roomi, Muhammad Amir Zia, Amir Ijaz

Abstract


Potato is a very important food crop and is adversely affected by fungus. Agrobacterium-mediated transformation can play an important role in the improvement of potato. The present study was conducted to optimize the different factors affecting Agrobacterium-mediated transformation of chitinase gene. Nodes were used as explant of potato cultivars Desiree and Sh-5. Agrobacterium tumefacien strain LBA4404 harboring a pB1333-EN4-RCG3 plasmid having chitinase gene and selectable marker hygromycin gene under the control of the CaMV 35S promoter was used. The parameters optimized for the potato transformation includes co-cultivation time, cefotaxime concentration, and days to pre-selection. The results show that the infection time (2 min) gave best mean value of transformation efficiency, that is, 2.9 and 2.1 in Desiree and Sh-5, respectively. The over growth of Agrobacterium were controlled with a concentration of 200 mg/l of cefotaxime and obtained maximum mean value of transformation efficiency of 3.38 and 3.10 in Desiree and Sh-5, respectively. The pre-selection period seven days prior to selection were considered effective for regeneration of explants and high transformation efficiency. The high mean value of regeneration (3.08 and 2.82) and transformation efficiency (3.00 and 2.60) was observed in Desiree and Sh-5, respectively. The putative transgenic plants were analyzed through PCR by using RCG-3 specific primers.

Key words: Agrobacterium, potato, transformation, rice chitinase gene, factors.




http://dx.doi.org/10.5897/AJB11.1961
AJOL African Journals Online