Transgenic resistance by N gene of a Peanut bud necrosis virus isolate of characteristic phylogeny.

The nucleocapsid protein (N) gene of a Tospovirus devastating tomato crop in the south Indian state of Tamil Nadu was cloned and characterized. The high identity of the cloned sequence to a Peanut bud necrosis virus (PBNV) tomato isolate (97.8/99.6% nucleotide/amino acid) and a PBNV peanut isolate (94.4/96.3% nucleotide/amino acid) identified the Tospovirus as an isolate of PBNV, designated PBNV Coimbatore tomato (PBNV CT) isolate. Phylogenetic analysis of PBNV CT N gene provided useful insights into the movement and evolution of PBNV within Indian Territory. The characteristic phylogeny of PBNV CT N gene implied its potential to be an efficient transgene to confer effective PBNV resistance on crop plants. The efficacy of PBNV CT N gene in conferring PBNV resistance was studied by generating tobacco (Nicotiana tabacum L. cv Wisconsin) lines transgenic to the sense or antisense version of the gene. Several transgenic lines showed transgenic mRNA and/or protein accumulation, ranging from very high to undetectable levels, accompanied by different degrees of PBNV resistance. The undetectable or very low levels of transgene transcripts in certain PBNV-resistant sense or antisense N gene transgenic lines suggested RNA-mediated resistance by post-transcriptional gene silencing (PTGS) mechanism. However, PBNV resistance of certain transgenic lines with high levels of N gene transcripts was suggestive of possible operation of RNA-mediated non-PTGS mechanism(s) of resistance in those lines. Moreover, the high levels of N protein in certain PBNV-resistant sense N gene transgenic lines suggested protein-mediated resistance. The results predict the potential of PBNV CT N gene to confer effective PBNV resistance on tomato and other economically important crops.

Potential threat of a new pathotype of Papaya leaf distortion mosaic virus infecting transgenic papaya resistant to Papaya ringspot virus.

A virus identified as a new pathotype of Papaya leaf distortion mosaic virus (PLDMV, P-TW-WF) was isolated from diseased papaya in an isolated test-field in central Taiwan, where transgenic papaya lines resistant to Papaya ringspot virus (PRSV) were evaluated. The infected plants displayed severe mosaic, distortion and shoe-stringing on leaves; stunting in apex; and water-soaking on petioles and stems. This virus, which did not react in enzyme-linked immunosorbent assay with the antiserum to the PRSV coat protein, infected only papaya, but not the other 18 plant species tested. Virions studied under electron microscope exhibited morphology and dimensions of potyvirus particles. Reverse transcription-polymerase chain reaction conducted using potyvirus-specific primers generated a 1,927-nucleotide product corresponding to the 3' region of a potyvirus, showing high sequence identity to the CP gene and 3' noncoding region of PLDMV. Search for similar isolates with the antiserum against CP of P-TW-WF revealed scattered occurrence of PLDMV in Taiwan. Phylogenetic analysis of PLDMV isolates of Taiwan and Japan indicated that the Taiwan isolates belong to a separate genetic cluster. Since all the Taiwan isolates infected only papaya, unlike the cucurbit-infecting Japanese P type isolates, the Taiwan isolates are considered a new pathotype of PLDMV. Susceptibility of all our PRSV-resistant transgenic papaya lines to PLDMV indicates that the virus is an emerging threat for the application of PRSV-resistant transgenic papaya in Taiwan and elsewhere.

Pyramiding transgenic resistance in elite indica rice cultivars against the sheath blight and bacterial blight.

Elite indica rice cultivars were cotransformed with genes expressing a rice chitinase (chi11) and a thaumatin-like protein (tlp) conferring resistance to fungal pathogens and a serine-threonine kinase (Xa21) conferring bacterial blight resistance, through particle bombardment, with a view to pyramiding sheath blight and bacterial blight resistance. Molecular analyses of putative transgenic lines by polymerase chain reaction, Southern Blot hybridization, and Western Blotting revealed stable integration and expression of the transgenes in a few independent transgenic lines. Progeny analyses showed the stable inheritance of transgenes to their progeny. Coexpression of chitinase and thaumatin-like protein in the progenies of a transgenic Pusa Basmati1 line revealed an enhanced resistance to the sheath blight pathogen, Rhizoctonia solani, as compared to that in the lines expressing the individual genes. A transgenic Pusa Basmati1 line pyramided with chi11, tlp, and Xa21 showed an enhanced resistance to both sheath blight and bacterial blight.