Efficient genetic transformation of Sorghum using a visual screening marker

Area of Science:

• Plant Biotechnology
• Molecular Biology
• Agricultural Science

Background:

• Grain sorghum (Sorghum bicolor) transformation is crucial for crop improvement.
• Efficient methods are needed to introduce genes for enhanced disease and drought resistance.
• Visual markers simplify the selection of transgenic plants.

Purpose of the Study:

• To develop an efficient Agrobacterium-mediated transformation protocol for grain sorghum.
• To introduce genes for enhanced fungal disease and drought resistance using a visual reporter.
• To validate the use of green fluorescent protein (GFP) as a screening marker.

Main Methods:

• Agrobacterium-mediated transformation of immature sorghum embryos (Tx 430, C401, Pioneer 8505).
• Use of a binary vector containing the reporter gene (gfp) and target gene (tlp) under the maize ubi 1 promoter.
• Visual screening of calli and plants for GFP expression.
• Confirmation of transgene integration and expression using Southern and Western blot analyses.

Main Results:

• Successfully generated 1011 fertile transgenic sorghum plants from 2463 embryos.
• Demonstrated 100% correlation between GFP expression and the presence of the target tlp gene.
• Confirmed stable transgene integration and expression in selected transgenic lines and their progeny.
• Showed efficient segregation of transgenes and varying copy numbers in offspring.

Conclusions:

• Green fluorescent protein (GFP) serves as an effective visual marker for efficient sorghum transformation.
• This method enables the production of stably transformed sorghum plants resistant to fungal diseases and drought.
• The protocol bypasses the need for antibiotic or herbicide selection agents, offering a more streamlined approach.