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Open Access Research article

Agrobacterium-mediated transformation of rough lemon (Citrus jambhiri Lush) with yeast HAL2 gene

Shawkat Ali123*, Abdul Mannan14, Mohamed El Oirdi2, Abdul Waheed5 and Bushra Mirza1

Author Affiliations

1 Department of Biochemistry, Quaid-i-Azam University, Islamabad, Pakistan

2 Département de Biologie, Université de Sherbrooke, Québec, Canada

3 Horticulture R & D Centre Agriculture and Agri-Food Canada 430, Boulevard Gouin, St-Jean-sur-Richelieu, Québec, Canada

4 Department of Pharmaceutical Sciences, COMSATS Institute of Information Technology, Abbottabad, 22060, Pakistan

5 Department of Botany, University of Arid Agriculture, Rawalpindi, Pakistan

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BMC Research Notes 2012, 5:285  doi:10.1186/1756-0500-5-285

Published: 12 June 2012

Abstract

Background

Rough lemon (Citrus jambhiri Lush.) is the most commonly used Citrus rootstock in south Asia. It is extremely sensitive to salt stress that decreases the growth and yield of Citrus crops in many areas worldwide. Over expression of the yeast halotolerant gene (HAL2) results in increasing the level of salt tolerance in transgenic plants.

Results

Transformation of rough lemon was carried out by using Agrobacterium tumefaciens strains LBA4404 harboring plasmid pJRM17. Transgenic shoots were selected on kanamycin 100 mg L-1 along with 250 mg L-1 each of cefotaxime and vancomycin for effective inhibition of Agrobacterium growth. The Murashige and Skoog (MS) medium containing 200 μM acetoseryngone (AS) proved to be the best inoculation and co-cultivation medium for transformation. MS medium supplemented with 3 mg L-1 of 6-benzylaminopurine (BA) showed maximum regeneration efficiency of the transformed explants. The final selection of the transformed plants was made on the basis of PCR and Southern blot analysis.

Conclusion

Rough lemon has been successfully transformed via Agrobacterium tumefaciens with β-glucuronidase (GUS) and HAL2. Various factors affecting gene transformation and regeneration efficiency were also investigated.

Keywords:
Genetic transformation; Yeast halotolerant gene (HAL2); Citrus jambhiri Lush and Agrobacterium tumefaciens