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Evaluation of Brassica juncea (L.) Czern x B. rapa/B. carinata hybrids and their advanced progenies against two major fungal diseases: molecular, morphological and biochemical characterization

Student Name: Ms Kadambari Gupta
Guide: Dr Abha Agnihotri
Year of completion: 2006

Abstract:

The present research work was undertaken during 2001 –2005 at the Centre for Bioresources and Biotechnology, TERI School of Advanced Studies, New Delhi to transfer resistance/ tolerance to white rust and alternaria blight from Brassica rapaand Brassicacarinatato Brassica junceathrough in vitroembryo rescue as well as in vivo seed set. Interspecific hybrids of B. juncea× B. rapa/B. carinata were grown and characterized using morphological and molecular markers to assess the variability for both the diseases. The selected plant progenies were advanced and screened for agro-morphological traits and biochemical parameters with varying degree of diseases response. Two genotypes RESBR 219 and RESBR 350 of B. raparesistant to Peronospora parasiticaand Albugo candidarace 2 and var. Kiran of B. carinataresistant to white rust and tolerant to alternaria blight were selected. The B. junceagenotypes selected for resistance to P. parasiticaand A. candidarace 7 were RESBJ 830 and RESBJ 837, while TERI (OE) M21-1 was selected for improved nutritional oil quality profile (low erucic acid and high oleic acid). The salient findings of the present research work are summarized as follows:

Putative hybrids were obtained through in vivo(4.7%) and in vitro(2.4%) techniques with the majority of hybrids being produced through ovule culture. Post fertilization barriers existed in in vivoseed germination and required an in vitrophase for establishment of hybrid seedlings. More hybrids were obtained with the amphidiploid (6.5%) species as male donor than diploid species (2.1%). ISSR markers confirmed the hybrid nature of the plantlets. Morphologically hybrids were intermediate to their parent genotypes but with male donor identifying features in the B. junceax B. carinata cross.

The parent genotypes of B. junceaand B. rapashowed a differential response to A. candidaisolates; with the former being resistant to field isolate of A. candida derived from B. rapaand the latter moderately resistant to B. junceafield isolates. Brassica junceavar. Varuna, parent genotype TERI (OE) M21-1 as well as B. rapavar. Pusa Gold were susceptible or moderately susceptible to all the isolates, whereas B. carinatagenotype Kiran was resistant to all the field isolates under controlled epiphytotic conditions. Brassica junceaisolate collected at Delhi (AcBjD) was more virulent as compared to the Pantnagar isolate (AcBjP). Overall, host-pathogen response was greater using the mixture of isolates than 8 the individual isolates. Both alternaria blight fungal toxin and pathogen suspensions distinguished between B. junceaand B. carinatacategorizing them as susceptible and resistant respectively. However, different B. junceagenotypes gave differential disease response when inoculated with conidialsuspension but showed a similar moderately resistant disease reaction oninteraction with crude fungal toxin of alternaria blight.

F2and BC1plants of the cross B. junceax B. rapaproduced 67.6% and 80.0% B. junceatype plants while all plants in F3and F2BC1, BC1F2and BC2generations resembled the female parent with partial characteristic of the male donor evident in F3and BC1F2. Hybrids from the cross B. junceax B. rapawere resistant to white rust (DI<1.0) but differed in their disease response in subsequent generations. F3and F2BC1plants showed no visible disease symptoms. DI increased from BC1(1.53) to BC1F2(1.86) and BC2(2.48) in the cross combination involving parent genotype RESBJ 830, while there were no differences in DI of BC1(1.41), BC1F2(1.28) and BC2 (1.75) plants in crosses involving RESBJ 837.The resistance was either equal to the male donor or superior to both the parents. Overall, a higher number of resistant plants were obtained from the cross combination RESBJ 837 x RESBR 219 than the cross RESBJ 830 x RESBR 350. A total of 76 resistant plants (DI≤1.0)having B. junceatype morphological characteristics were selected from the F3and F2BC1, BC1F2and BC2generations. A positive correlation was observed between growth stages of plants and white rust disease infestation in all generations as the disease progressed from 75 to 90 DAS (r=0.93) and from 90 to 110 DAS (r>0.97) under field conditions.

F2and BC1populations of B. junceax B. carinatacross had 72% intermediate and 60% B. junceatype plants.The plants from cross combination involving RESBJ 837 and TERI (OE) M21-1 were B. junceatype in BC1F2and BC2.On the other hand, plants from the cross combination RESBJ 830 showed intermediate morphology in BC1F2and B. junceatype BC2 plants with strong male donor characteristics like short main branch. A characteristic red dot present at the tip of anthers on hybrids was observed on F2plants. The red dot presence diminished with successive advanced generations (BC1, BC2and BC1F2). The presence of purple colour inherited from the male donor was observed in all the generations.Plant progenies with TERI (OE) M21-1 as female parent were brown seeded in all generations, whereas plant progenies involving RESBJ 837 and RESBJ 830 segregated into yellow and brown seeded plants in the ratio of 10: 19 in BC1, 5: 42 in BC1F2 and 10:7 in BC2.

Hybrids, F2and BC1plantsfrom the cross B. junceax B. carinatashowed only a few visible disease symptoms for white rust. The BC1 plants from all three cross combinations were tolerant to alternaria blight at both the mature leaf growth stage and the pod formation stage. An equivalent disease response was observed in plants derived from cross combinations involving the female genotypes RESBJ 830 (0.81) and RESBJ 837 (0.94). These were superior to the plants obtained from cross combination involving TERI (OE) M21-1 (1.47) as female parent. All the BC1 plants exhibited a higher resistance than their respective female parents (>3.4). The BC2plants from these three different B. junceagenotypes did not show variation in average disease response at the mature leaf growth stage and were resistant to white rust (average DI <1.0) and alternaria blight (DI<2.0).

In all the generations, the proportion of plants resistant to white rust was higher (79.16-100%) and consistent as compared to the varying proportions of the populations resistant to alternaria blight (7.50% in F2to 100% in BC2). Overall, plants with the female genotype RESBJ 837 exhibited a higher resistance to both white rust and alternaria blight as compared to the plants with the genotype TERI (OE) M21-1K followed by RESBJ 830. A total of 69 BC1F2and BC2B. junceatype plants were selected with resistance to white rust (DI≤1.0)and alternaria blight (DI<2.0).The BC1F2and BC2plants obtained from two different cross combinations of the cross B. junceax B. rapadid not differ for important yield traits and tended to resemble the female parent. The F2BC1and BC2were equivalent to the female parent for important yield attributing traits including 1000 seed weight. The fatty acid profile of seeds of these plants resembled their female parent with high erucic and low oleic acid content but one plant from each of the BC1F2and BC2populations showed moderately low glucosinolate content of 45.55 μmol/g meal.

The BC1F2plants from cross B. junceax B. carinataresembled either the male donor (RESBJ 830 x Kiran) or were intermediate between the parents (RESBJ 837/ TERI (OE) M21-1 x Kiran). The BC2progenies were equivalent to and resembled the female parent for >50% of the agronomic traits, including days to maturity, number of pods on main shoot and 1000 seed weight. The exception was seeds per pod which was highest in the cross involving RESBJ 837 (16 seeds) followed by TERI (OE) M21-1 (14 seeds) and RESBJ 830 (6 seeds). The fatty acid profile and glucosinolate content of plants involving RESBJ 830 and RESBJ 837 was similar to their female parents with high erucic and low oleic acid content as well as high glucosinolate content, while significant desirable variation was observed in the plants from TERI (OE) M21-1 in all generations.

The F2plants obtained from TERI (OE) M21-1 showed desirable low glucosinolate content (<30 μmol/g meal) in three plants and moderately low in 12 plants (<60 μmol/g meal) (42.8%). The fatty acid profile in seeds of BC1disease tolerant plants indicated a transition towards the female parent withthree plants exhibiting high oleic acid (40.0 to 43.0%) and low erucic acid (1.2 to 5.0%) content. An unexpected, low glucosinolate content in seeds of three plants (13.5 to 29.0 μmol/g meal) and moderately low levels in seven plants (30-60 μmol/g meal) were identified along with tolerance to white rust (DI<1.0). A white rust tolerant BC1plant with unusually high levels of palmitic (17.48%) and stearic acid (10.62%), along with moderately low oleic (15.0%) and erucic acid (25.0%) as well as low glucosinolate content of 29.04 μmol/g meal was also identified.The fatty acid profile of BC1F2and BC2progenies of this cross resembled the female parent with low erucic (< 1.0%) and high oleic acid content (> 44.0%) along with tolerance to white rust and alternaria blight. No correlation was observed between erucic acid and the disease response to white rust (r=0.21) or alternaria blight (r=0.19) nor between glucosinolate content and the response of plants to white rust (r=0.19) and alternaria blight (r=0.22).

It is concluded that hybrids can be obtained from B. junceax B. rapa/ B. carinataunder in vivoconditions but require an in vitrophase for seedling establishment. The ISSRs utilized for studying hybrid nature indicated the applicability of ISSR markers inparentage studies as an efficient tool for screening wide hybrids inBrassica species. To the best of our knowledge, till date no published report is available on this aspect. On the whole, 76 B. junceatype plants were selected for white rust resistance (DI ≤1.0; less than 5% leaf area covered with white rust pustules) from advanced generations of the cross B. junceax B. rapaalong with resistance to Peronospora parasiticaunder natural field conditions. Similarly, 69 B. junceatype plants were selected for both white rust (DI<1.0) and alternaria blight resistance (DI < 2.0; less than10% leaf area covered with alternaria blight symptoms) from the crossB. juncea xB. carinata.

The important outcomes of the present research were identification of B. junceatype plants with:
1) White rust resistant/ tolerant B. junceaplants from two different resistant sources of white rust, B. rapaand B. carinata
2) Combined resistance/ tolerance in B. junceaplants to both white rust and alternaria blight from B. carinata.
3) High tolerance to both the fungal diseases combined with yellow seed coat colour in B. juncea

Nutritionally desirable fatty acid profile (low erucic and high oleic acid content) along with disease tolerance to white rust and alternaria blight in B. juncea. Since incorporation of resistance to fungal diseases is economically and environmentally important for increasing productivity, besides minimizing the use of fungicides, these novel elite genotypes with adequate disease resistance traits and nutritionally desirable oil quality profile identified in the present study can be utilized for developing nutritionally improved quality B. junceacultivars with white rust and alternaria blight resistance, for nutritional security in an environment friendly manner.

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