DEVELOPMENT OF INTERSPECIFIC HYBRIDS IN Trifolium THROUGH EMBRYO RESCUE AND THEIR CHARACTERIZATION FOR STEM ROT TOLERANCE
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Date
2012
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Punjab Agricultural University, Ludhiana
Abstract
The present investigation entitled, “Development of interspecific hybrids in
Trifolium through embryo rescue and their characterization for stem rot tolerance” was
carried out using Trifolium alexandrinum (cv.s Wardan and BL1) as female parent and T.
vesiculosum as male parent. A total of 1053 florets of Wardan were pollinated (AV), from
which only 27 embryos were rescued and 12 embryos germinated in vitro on EC medium (MS
+ 0.5 mgl-1 BAP + 3.0% sucrose). In BL1 × T. vesiculosum (BV) cross, out of 1128 florets
pollinated, 69 embryos were rescued and only 39 of them germinated in vitro. The maximum
embryo survival was seen in the florets collected at 11-12 DAP and the embryos at
cotyledonary stage responded best to the culture conditions. A total of 12 putative AV and 39
putative BV hybrids were obtained. Clonal multiplication was done on LSP3 medium (L2 +
0.0008 mgl-1 NAA + 0.15 mgl-1 BAP + 2.5% sucrose) and 99.98% shoot regeneration was
observed. Root induction (16.67%) in shoots was observed on RL2 medium (RL + 0.42 mgl-1
IAA + 2.5% sucrose). Molecular characterization of the putative hybrids was done using 23
SSR markers, out of which 8 were found to be polymorphic and confirmed one AV hybrid
(AV1) and three BV hybrids (BV3, BV4 and BV12). Nine clones of AV1 hybrid and 12
clones of BV3 hybrid along with 5 clones of each parent were transplanted in the polythene
bags containing autoclaved soil. Morphological data was recorded on two month-old parents
and the hybrids. The AV hybrid was significantly taller than both the parents. Likewise BV
hybrid was also significantly different from both the parents in plant height. The hybrids were
further screened for resistance against stem rot by inoculating the collar regions with
Sclerotinia trifoliorum culture raised on whole gram. Observations made after 14 days of
inoculation of fungal culture showed 11.11% infection in AV1 and 8.33% infection in case of
BV3 hybrids. Thus, hybrid AV1 was moderately susceptible and BV3 hybrid was moderately
resistant to the disease.
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