**10. Recommended strategies for improvements**

There is adequate evidence gathered in this study indicating that, the soybean genotypes used will variably express and transmit the transgenes if any event of transformation is to take place. This was clearly demonstrated by the varied responses formed in plant cell initiation and microshoots formation during callus induction and shoot multiplication. Moreover, the varied intensities in oxidative browning of tissues, chlorosis and subsequent wounded tissue necrosis exhibited by explant from different genotypes, indicated the extend of the problems associated with *Agrobacterium*-mediated genetic transformation of legumes. For the successful optimisation of protocol routinely used for transformation of a wide range of soybean varieties, this tool requires the following amendments:

**11. Conclusions**

**Acknowledgements**

(nGAP), South Africa.

**Thanks**

their support.

**Author details**

Phetole Mangena

Sovenga, South Africa

**References**

The use of double cotyledonary-node explants still remain superior for establishment of soybean cultures–callus and shoot proliferation. This is so, because the efficiency of the cultures established relied primarily on the explant type, as one of the culture factors. Explant amenability to *Agrobacterium* infection, antibiotics and growth regulator regimes impacted highly on the culture successes observed. A successful shoot and callus induction in the control media was illustrated. The negative impacts of *Agrobacterium* infection and effect of antibiotics on the culture was also observed. However, these findings clearly demonstrate that, more work still need to done, focusing on the optimisation of tissue culture conditions and bacterial cultures. For the main purpose of developing a high frequency, genotype independent and efficient

The Role of Plant Genotype, Culture Medium and *Agrobacterium* on Soybean Plantlets…

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I would like to sincerely acknowledge the financial support by the Department of Higher Education and Training (DHET), under the New Generation of Academics Programme

My sincere gratitude and thanks goes to Dr. P.W. Mokwala and Prof R.V. Nikolova for their support and mentorship. Their words of encouragement will continue ringing in my ears. Many thanks to my family, friends, colleagues and the entire Department of Biodiversity for

Department of Biodiversity, School of Molecular and Life Sciences, University of Limpopo,

[1] Messina MJ. Legumes and soybeans: Overview of their nutritional profiles and health

effects. American Society for Clinical Nutrition. 1999;**70**(3):439-450

Address all correspondence to: mangena.phetole@gmail.com

protocol for use in *Agrobacterium*-mediated genetic transformation of soybean.


Other affordable method such as *in-planta Agro*-injection must be considered by laboratories to generate new genetically improved soybean plants. This technique was introduced by Chee et al. [65] in genetic transformation of soybean and kidney bean by *Agro*-injecting seeds with a suspension of *Agrobacterium* strain EHA101 with pIG121 vector plasmid containing genes for neomycin phosphotransferase (NPTII), hygromycin phosphotransferase (HPT) and β-glucuronidase (GUS). The soybean and kidney bean seeds were surface sterilised using 0.6% sodium hypochlorite and germinated on moistened sterile paper towels at 25°C for 24 h in darkness. *Agro*-infection yielded 12% transgenic soybeans and 24% of transgenic kidney beans identified using NPTII amplified by polymerase chain reaction (PCR). Generally, there are difficulties in the *in vitro* regeneration and selection of transgenic plants during *Agrobacterium*-mediated genetic transformation. But, this technique is undoubtedly the best tool available for the transfer and expression of transfer DNA in host plant cells.
