Transgenic rice lines constitutively co-expressing tlp-D 34 and chi 11 display enhancement of sheath blight resistance
Biologia Plantarum(2012)
摘要
Transgenic rice ( Oryza sativa L. subsp. indica cv. White Ponni) constitutively expressing the rice thaumatin-like protein gene ( tlp-D 34, PR-5) individually or in combination with the rice chitinase gene ( chi 11, PR-3) was generated using an Agrobacterium vir helper strain with multiple copies of pTiBo542 vir B and vir G. Transformation with the tlp-D 34 gene alone and tlp-D 34 + chi 11 genes yielded five and seven single-copy transgenic lines, respectively. Southern blot analysis with two probes, one flanking the right T-DNA border and the second flanking the left T-DNA border, confirmed that all transgenic plants harboured single and complete T-DNA copies. Homozygous transgenic lines were first identified in the T 1 generation by Southern blot analysis and were subsequently confirmed by segregation analysis of T 2 plants. Accumulation of transcripts encoded by the transgenes was confirmed in T0 plants and homozygous T 2 plants by Northern blot analysis. The homozygous T2 plants harbouring tlp-D 34 + chi 11 genes showed 2.8- to 4.2-fold higher chitinase activity. Western blot analysis revealed the accumulation of thaumatin-like protein and chitinase in the respective transgenic plants. Upon infection with Rhizoctonia solani , the disease index reduced from 100 % in control plants to 65 % in a T 3 homozygous transgenic line T4 expressing the tlp-D 34 gene alone. In a T2 homozygous transgenic line CT22 co-expressing tlp-D 34 and chi 11 genes, the disease index reduced to 39 %.
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关键词
Agrobacterium tumefaciens, homozygous transgenic lines, Oryza sativa, pathogenesis-related protein, Rhizoctonia solani, fungal resistance
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