Screening of mungbean and ricebean entries for Mungbean Yellow Mosaic Virus Resistance through Agroinfection Department of Plant Molecular Biology and Biotechnology, Centre for Plant Molecular Biology, Tamil Nadu Agricultural University, Coimbatore. M. Sudha, Research Scholar

- Dr. Norman Borlaug

White flies Yellow mosaic disease in pulses

 Agroinfection is an effective method by which infectious viral clones are introduced into plants using Agrobacterium tumefaciens  A partial tandem repeat of a virus is constructed and placed in the T-DNA of a binary vector and introduced into A. tumefaciens  Upon inoculation of plants with the Agrobacterium strains and transfer of the T-DNA, the viral genome is released in the plant cells from the partial tandem repeat of the virus Agroinfection

 Infectivity analysis of two variable DNA B components of Mungbean yellow mosaic virus- Vigna in Vigna mungo and Vigna radiata  Mungbean yellow mosaic virus- Vi Agroinfection by codelivery of DNA A and DNA B from one Agrobacterium Strain  Analysis of an isolate of Mungbean yellow mosaic virus (MYMV) with a highly variable DNA B component  Infectivity analysis of a soybean isolate of Mungbean yellow mosaic India virus by agroinoculation Studies on Agroinfection in pulses

Agrobacterium strains  VA221 - (KA 22 DNA B + KA30 DNA A)  VA239 - (KA 27DNA B + KA30 DNA A) These strains cloned from infected black gram were obtained from Dr. K. Veluthambi (Madurai Kamaraj University)

MYMV Symptom expression in Co 5 black gram for VA 221 and VA 239 strains

Susceptible reaction to MYMV through agroinfection in mungbean LM 122 (field susceptible)

Field screening of germplasm entries against MYMV S.NoGermplasmNumber of entries screened Number of resistant entries 1.Mungbean Ricebean24

Agroinfection screening of mungbean entries against MYMV Reaction Number of entries Number of entries Resistant to VA2213 Resistant to VA 2391 Susceptible24 Total28

S.NoEntriesS.NoEntriesS.NoEntries 1KMG 14911ML 13121PDM PUSA ML COGG 917 3PUSA 9030-BOK13EC KMG 189 4SEL 214ZKM 2624SP 84 5PLS VGG 2425SU PLS 303-B16SU VGG 148 7VBM-92-3-B17SU VGG NL 518SU ML PDM VGG COGG-92420ML 818 Resistant to VA221 strain Resistant to VA239 strain List of field resistant mungbean entries subjected to agroinfection

MYMV Symptom(s) expression in mungbean – ML 1108 (VA221 resistant )

MYMV Symptom(s) expression in mungbean - ML 818 (VA239 resistant )

FOR VA221 Strain 703 bp PCR amplification in MYMV agroinoculated mungbean entries FOR VA239 Strain Positive control 1-20 : Agroinoculated entries Primer used – Coat protein region of DNA A 100bp CO 5 blackgram-VA221 2.CO5 blackgram-VA bp

Agroinfection screening of ricebean entries against MYMV ReactionNumber of entries Resistant24 Susceptible 0 Total24

S.NoEntriesS.NoEntries 1V. umbellata TNAU red13LRB 89 green 2LRB 4114LRB 11 3LRB 29715LRB 29 l.green mottled 4LRB 110 Yellow16LRB 35 5LRB 113 original17TNAU Yellow 6LRB 2318LRB 111 7V. Umbellata TNAU green19LRB 104 8LRB 113 black20LRB 282 9LRB mottled green21LRB 5 mottled 10V. Umbellata dark green22LRB 55 11LRB tendrills23 LRB 109 green 12LRB 15 green24 LRB 15 green List of Rice bean entries subjected to agroinfection

Resistance reaction to MYMV through agroinfection in ricebean LRB 297

Resistant symptom expression in Ricebean TNAU RED along with positive check blackgram Co 5 through agroinfection

1000bp bp Lane1 : Uninoculated Control TNAU red Lane 2 : First day Lane 3 : Second day Lane 4 : Nineth day Lane 5 : Tenth day Lane 6 : Twelth day Lane 7 : Fifteenth day Lane 8 : Seventeenth day Lane 9 :Twenty-second day Lane10 : Immediately after inoculation( embryos alone) Lane11 : Third day ( embryos alone) Lane12 : Fourth day (embryos alone) Lane13 : Nineth day Lane14 : Tenth day Lane15 :Twelth day Lane16 : Fifteenth day Lane17 : Seventeenth day Lane 18 : Eighteenth day Lane19 : Twentith day Lane 20 : Twenty-first da y PCR amplification of agroinoculated ricebean TNAU RED for VA221 strain 700 bp 800 bp

Lane 1 : 2 month old lower leaves Lane 2 : 1 month old upper leaves Lane 3 : 1 month old lower leaves Lane 4 : Twenty four days old leaves Lane 5 : Fifteenth days old leaves 100bp bp 700 bp 800 bp PCR amplification of agroinoculated ricebean TNAU RED VA221 strain at different growth stages

Ongoing works…  Real-Time PCR assays for detection and quantification of MYMV viruses for the identified resistant mungbean entries which were strain specific  Identification of resistant proven donors for MYMV resistance through agroinfection which can be used as a genetic material