Showing papers in "Journal of Plant Biochemistry and Biotechnology in 1998"
TL;DR: This review focuses on the genes for the enzymes 5-enolpyruvyl-3-phosphoshikimlc acid synthase and the glyphosate oxidoreductase, used to genetically engineer plants that are resistant to the herbicide glyphosate.
Abstract: This review focuses on the genes for the enzymes 5-enolpyruvyl-3-phosphoshikimlc acid synthase (EPSPS) and the glyphosate oxidoreductase (GOX). These genes have been used to genetically engineer plants that are resistant to the herbicide glyphosate. Overproduction of glyphosate-insensitive.EPSPS in transgenic crops has been used to overcome the deleterious effuts of this herbicide. The introduction into plants of GOX also confers glyphosate tolerance to plants and augments the tolerance of transgenic plants already expressing a glyphosate tolerant EPSPS. These genes also provide a method for selecting transformed plant tissue using the glyphosate tolerance as the selectable marker in the presence of inhibitory concentrations of glypllosate. Glyphosate tolerant transgenic plants of beet, corn, cotton, lettuce, poplar, potato, rapeseed. soybean, tobacco, tomato, and wheat have already been field tested and are entering agriculture.
38 citations
TL;DR: Polyclonal antisera were produced in albino white rabbits against intact teliospores of Karnal bunt (Tilletia indica) and the Immunoprobe generated was used for the development of Immunoblot binding assay for detecting Karnalbunt (KB) infections in wheat seed samples.
Abstract: Polyclonal antisera were produced in albino white rabbits against intact teliospores of Karnal bunt (Tilletia indica). The Immunoprobe generated was used for the development of Immunoblot binding assay for detecting Karnal bunt (KB) infections in wheat seed samples. The antiserum reacted strongly with intact teliospores of T. indica, Pantnagar isolate in agglutination reaction. Wheat grains with different grades of infection could be readily detected by Seed Immunoblot Binding Assay (SIBA). Karnal bunt infected wheat seeds when kept for vigour testing on nitrocellulose paper, formed a coloured imprint after the paper was immunoprocessed. The SIBA would not only be a better Indication of teliospores load on seed but also quality of seed in terms of vigour. This method is expected to be useful in routine monitoring of wheat lots for the presence of KB teliospores.
25 citations
TL;DR: Callus induced from immature embryos of wheat cv Kharchia 65 on Murashige and Skoog’s medium containing 2.5 mg l1 2,4-D was maintained In the regenerable state by subculturing every 5-6 weeks on medium supplemented with 2, 4-D and proline to maintain morphogenic competence for over four years.
Abstract: Callus induced from immature embryos of wheat cv Kharchia 65 on Murashige and Skoog’s medium containing 2.5 mg l1 2,4-D was maintained In the regenerable state by subculturing every 5-6 weeks on medium supplemented with 2,4-D (2.5 mg l-1) and proline (10 mg l-1). Addition of proline helped maintain morphogenic competence for over four years. The regenerating callus was analysed histologically about one year after first induction. Both somatic embryogenesis and shoot organogenesis were seen in the same callus tissue that contained typical stages of somatic embryoid development and evidence for the de novo shoot bud formation.
16 citations
TL;DR: Rhizospheric application of naringenin partially alleviated the deleterious effect of low temperature on nodulation status and nodule efficiency in Rhizobium leguminosarum.
Abstract: The production of Tsr factor by Rhizobium leguminosarum bv. viciae was influenced by low temperature (10°C) In the presence of seed exudate collected at 10°C and 25°C or naringenin (10fuM). Root exudate collected at 25°C and naringenin induced Tsr factor in R. leguminosarum causing thick and short root phenotype and root hair curling and deformation of host root. Root exudate collected at 10°C also induced root hair curling but Tsr activity was low. low temperature grown plants had poor nodulation, nitrogen fixation, nitrogen content and total blomass as compared to plants grown at 25°C. Rhizospheric application of naringenin partially alleviated the deleterious effect of low temperature on nodulation status and nodule efficiency.
15 citations
TL;DR: Reciprocal cross hybridization between DNA-A and ONA-B molecules showed that MYMV genome contains certain regions common to each other and the homology between ICMV andMYMV-Bg coat protein gene appeared to be less than 50%.
Abstract: Reciprocal cross hybridization between DNA-A and ONA-B molecules showed that MYMV genome contains certain regions common to each other. “Common region” of MYMV belongs to the 0.55 kb Kpnl-Clal fragment of DNA-A and 0.25 kb Xbal-Hind III fragment of DNA-B. Hybridization of DNA-A restriction fragments with the full length coat protein region probe to Indian cassava mosaic virus (ICMV) showed the location of MYMV coat protein coding region in DNA-A. The greater extent of homology of ICMV coat protein region falls within the 400 base pair region of Clal-Clal fragment in DNA-A of MYMV-Bg. The homology between ICMV and MYMV-Bg coat protein gene appeared to be less than 50%. Cross hybridization between DNA-A and DNA-B molecules helped to trim out the homologous region and the DNA-A and DNA-B specific probes could be tailored.
11 citations
TL;DR: Nine PAL genomic clones from tobacco have been isolated andGene specific probes based on the low homology region in the 5′ upstream region were used to study their induction patterns with MJ and elicitor and found that both MJ and elicit Induced the gPAL A transcription but with different time courses.
Abstract: Nine PAL genomic clones from tobacco have been isolated. Restriction map analysis of these clones showed that they can be divided into two groups, gPAL A and gPAL B. Both of the genes were further subcloned and sequenced. They showed a high homology for nucleotide and deduced amino acid sequences in the coding region. gPAL B showed closer ties with other solanaceous members than to gPAL A. Gene specific probes based on the low homology region in the 5′ upstream region were used to study their induction patterns with MJ and elicitor. It was found that both MJ and elicitor Induced the gPAL A transcription but with different time courses. PAL A mRNA level in elicitor-treated cells peaked after 4 h of the treatment whereas in the MJ-treated cells PAL A mRNA peaked after 6-8 h of the treatment. Neither MJ nor elicitor activated the gPAL B transcription.
10 citations
TL;DR: Of the four different cytokinins tested, BA was most effective in inducing multiple shoot buds in the explants, and was recorded on Murashige and Skoog’s medium supplemented with 2.0 mg/l BA.
Abstract: An efficient protocol is described for large scale in vitro propagation of east Indian rosewood (Dalbergia latifolia Roxb.) using cotyledonary nodes derived from axenic seedlings. Of the four different cytokinins tested, BA was most effective in inducing multiple shoot buds in the explants. High frequency shoot proliferation (93%) coupled with maximum number of shoot formation (10-12 shoots/explant) was recorded on Murashige and Skoog’s medium supplemented with 2.0 mg/l BA. The frequency of shoot proliferation declined at higher levels of cytokinins. Shoot culture was multiplied by subculturing the original cotyledonary node on a fresh shoot multiplication medium each time aHer excising the newly formed shoots. Shoots obtained from each passage were multiplied further as nodal explants and each node produced 3-4 shoots. In two months from a single cotyledonary node, about 90 shoots were obtained. Rooting was induced in 72% of the regenerated shoots on half-strength MS containing IAA, IBA and IPA each at 1.0 mg/l. Rooted plantlets were hardened off and eventually established in soil.
9 citations
TL;DR: The data suggest the existence of half-and-half distribution of sites which is a manifestation of a pre-exlstent site heterogeneity in the oligomeric protein molecule.
Abstract: Kinetics of urease-catalysed urea hydrolysis follows Arrhenius equation in the temperature range 10-50°C and shows an energy of activation equal to 7.14 kcal/mol. The kinetics of thermal inactivation of the enzyme is biphasic, In that half of the initial activity is destroyed more rapidly than the remaining half. The data are consistent with the rate equation: At = Afast·e-kfast-t + Aslow ·e-Kslow-t where At is the residual activity at time t, Afast and Aslow, kfast and kslow are the amplitudes and the first-order rate constants of the fast and the slow phases, respectively. A similar activity decay (namely blphaslc) is also observed on storing the enzyme at +4 and −4OC. The data suggest the existence of half-and-half distribution of sites which is a manifestation of a pre-exlstent site heterogeneity in the oligomeric protein molecule.
9 citations
TL;DR: The data suggests the existence of site-site heterogeneity in oligomeric urease molecule from pigeonpea and the time course of thermal inactivation can be described by the following equation, consisting of two first order terms: At = Afast + Aslow.
Abstract: The purified urease from pigeonpea was moderately stable at −10°C. The shelf-life of the enzyme on storage in 0.1 M Tris-acetate buffer, pH 6.5, at −10°C showed a single exponential decay with a t1/2 of approx. 30 days. In the presence of additives like 5mM dithiothreitol the t1/2 increased to 223 days at the same temperature, in a single exponential decay. The Arrhenius plot of the kinetics of the pigeonpea urease catalysed urea hydrolysis over the temperature range 27 to 77°C, was linear. The Q10 value was found to be 1.46. The energy of activation calculated from the Arrhenius equation was found to be 5.1 kcal/mole active site. The thermal denaturation of pigeopea urease at 65 and 70°C was found to obey biphasic kinetics in which half of the activity is destroyed faster than the remaining half. The time course of thermal inactivation can be described by the following equation, consisting of two first order terms: At = Afast.e-kfast + Aslow.e-kslow.t where, At is the residual activity at time t, Afast and Aslow, kfast and kslow are the amplitudes and the first-order rate constants of the fast and the slow phases, respectively. The data suggests the existence of site-site heterogeneity in oligomeric urease molecule from pigeonpea.
8 citations
TL;DR: Clonal propagation in vitro of raisin tree (Hovenia dulcis Thunb.) was achieved using axillary buds from mature trees and young plants and plantlets thus formed were successfully transplanted to the field after a short acclimatization period.
Abstract: Clonal propagation in vitro of raisin tree (Hovenia dulcis Thunb.) was achieved using axillary buds from mature trees and young plants. Explants cultured on Murashige-Skoog’s medium with 1/3 of the original salt concentration, supplemented with 0.5 mg l-1 BAP and 0.5 mg l-1 IAA, showed proliferation of new shoots in 4-5 weeks. Adventitious shoot proliferation was also stimulated in subsequent subcultures in the presence of BAP. The shoots rooted when transferred to 1/3 Murashlge and Skoog’s medium with 0.1 mg l-1 of IBA. Plantlets thus formed were successfully transplanted to the field after a short acclimatization period.
8 citations
TL;DR: Chromatographic studies performed with TLC showed the presence of hyoscyamine as the major alkaloid present in the callus tissues, differentiated shoots and roots of Hyoscyamus muticus, however, alkaloids content varied in different tissues.
Abstract: Alkaloid production has been observed in cotyledonary leaf derived callus tissues, and also in in vitro differentiated shoots, and roots of Hyoscyamus muticus. The callus tissue was developed form cotyledonary leaf explants on Murashige and Skoog medium enriched with 2 mg 1-1 2, 4-D and 0.5 mg 1-1 BAP. Cotyledonary leaf derived callus was proliferated in the same medium for 2 passages (1 passage 28-30 days). Green and compact callus was used for alkaloid extraction. Shoots and roots formed on MS medium containing 0.05 mg 1-1 NAA and 0.5 mg 1-1 BAP, and also compact, nodular and embryogenic calli from which these shoots and roots differentiated, were used for alkaloid extraction. Chromatographic studies performed with TLC showed the presence of hyoscyamine as the major alkaloid present in the callus tissues, differentiated shoots and roots. However, alkaloid content varied in different tissues. Differentiated roots were found to contain maximum amount of hyoscyamine.
TL;DR: Shoots can tolerate gradual depletion of sucrose upto a limit of 5 g l-1 under mixotrophic condition and inhibit shoot growth under photomixotrophic conditions.
Abstract: Clonally propagated shoots of teak (Tectona grandis Linn) were cultured in vitro under photomixotrophlc (sucrose 10-40 g l-1) and photoautotrophic (sucrose-free medium) conditions in MS medium containing kinetin and benzyl amino purine (0.1 mg l-1 each). Sucrose concentrations were gradually depleted in mixotrophic cultures. Growth and fresh weight of shoot chlorophyll a, b and total chlorophyll content of leaves were estimated. In sucrose-free medium, growth and chlorophyll synthesis decreased after limited period of 2-3 subcultures, whereas they got stimulated under photomixotrophic condition with 10-30 g l-1 sucrose; optimum being the medium with 30 g l-1 sucrose. Higher concentration of sucrose (40 g l-1) inhibited shoot growth. Shoots can tolerate gradual depletion of sucrose upto a limit of 5 g l-1 under mixotrophic condition.
TL;DR: A net influx of extracellular calcium in the differentiating cells is presumed to accompany the auxin-induced response and initial lack of polarity in the decapitated hypocotyl segments subjected to auxin treatment is discussed.
Abstract: Auxin-calcium interaction has been studied to understand their involvement in adventitious root initiation from the hypocotyl explants of sunflower (Helianthus annuus L.). When hypocotyl explants were cultured on MS medium (containing calcium), 1 mg l-1 IAA was found to be optimal for root induction. However, the hypocotyl explants washed in EGTA (10-5M) solution for the removal of extracellular calcium, when cultured on medium containing IAA and calcium, exhibited enhanced rooting response. When EGTA-washed explants were cultured on the medium supplemented with lanthanum chloride (10-6 and 10-5M), it resulted in the inhibition of the rooting response and this inhibitory effect could be alleviated by the simultaneous addition of IAA. Similar observations have been made by using calcium channel blockers, verapamil and TMB-8, and also a calmodulin inhibitor, trifluoperazine. A net influx of extracellular calcium in the differentiating cells is thus presumed to accompany the auxin-induced response. These results have been discussed in light of initial lack of polarity in the decapitated hypocotyl segments subjected to auxin treatment.
TL;DR: By using protein synthesis inhibitors it has been observed that light enhances translation of NR mRNA, which regulates supply of reductant via photosynthesis for activity of NR.
Abstract: Regulation of nitrate reductase (NR) by light is a complex process and is manifested through gene expression, both at the level of transcription and translation, covalent modification of the enzyme and supply of reductant. Light induces nia gene transcription by some still unknown mechanism. Light induction of nia mRNA is also mediated via phytochrome. By using protein synthesis inhibitors it has been observed that light enhances translation of NR mRNA. Light induction of NR synthesis follows the circadian rhythmicity. Cytokinin increases NR activity by regulating NR expression at the transcriptional level. In response to light-dark transition, NR is quickly inactivated/activated by phosphorylation/dephosphorylation. Inactivation of NR involves phosphorylation of the serine-543, located at the hinge 1 region connecting MoCo domain and cyt b domain of NR, by a Mg2+ dependent protein kinase and subsequent binding of an inhibitor protein. A type 2A protein phosphatase dephosphorylates/activates NR in response to light signal. Light also regulates supply of reductant via photosynthesis for activity of NR. Whether light is absolutely necessary for NR activity is also discussed.
TL;DR: Following cluster analysis based on similarity indices for the RFLP banding patterns observed, seven cytoplasmic groups within LSGP were identified and clearly indicates that besides serving as a source of diversity for agronomic and adaptation traits, broad-based gene pools can also provide diverse sources of cytopLasmic male sterility.
Abstract: Restriction fragment length polymorphism (RFLP) of mitochondrial (mt) DNA provides a rapid and effective method to assess heterogeneity among male sterile cytoplasms. Six isonuclear A-lines (81 A1, with Tift 23A1, cytoplasm, ICMA 88001 (= 81Av) with Violaceum cytoplasm, 81A (=81A4) with monodli = violaceum cytoplasm, Pb 310A2 and Pb 311A2 with A2 cytoplasm from L 66A, and Pb 406A3 with A3 cytoplasm from L 67A), nine cytoplasmic male-sterility sources from Large-Seeded Genepool (LSGP 6, LSGP 14, LSGP 17, LSGP 22, LSGP 28, LSGP 36, LSGP 43, LSGP 55 and LSGP 66) and two each from Early Genepool (EGP 33 and EGP 15) and Population Varieties (PV 1 and PV 2) were characterized for variation in their mitochondrial genomes following Southern blot hybridizations using homologous (pearl millet 13.6 kb, 10.9 kb, 9.7 kb and 4.7 kb clones) and heterologous (maize atp6 and coxl clones) mitochondrial DNA (mtDNA) probes. Following cluster analysis based on similarity indices for the RFLP banding patterns observed, we identified seven cytoplasmic groups within LSGP. Two (LSGP 43 and LSGP 66) of these were quite distinct from each other as well as from other cytoplasms. This clearly indicates that besides serving as a source of diversity for agronomic and adaptation traits, broad-based gene pools can also provide diverse sources of cytoplasmic male sterility. These new CMS sources were also compared with standard CMS systems and cytoplasm-specific restriction fragments were identified.
TL;DR: Polymorphism for peroxidase, esterase and acid phosphatase Isozymes in pollen grains of major Indian maize land races known as ‘Sikkim primitives’ were studied and revealed similarities in primitiveIndian maize landraces from Northeastern Himalayan region and Nal-Tel an ancient indigenous race of Mexico.
Abstract: Polymorphism for peroxidase, esterase and acid phosphatase Isozymes in pollen grains of major Indian maize land races known as ‘Sikkim primitives’ were studied and compared with primitive races of maize of Mexican origin and five species of teosinte, Coix lacryma-jobi, Chionachne koenigii and Sorghum bicolor. The isozyme patterns and the resulting dendrograms revealed similarities in primitive Indian maize landraces from Northeastern Himalayan region and Nal-Tel an ancient indigenous race of Mexico. Further, the Asiatic taxa of Maydeae, C. lacryma-jobi and Ch. koenigii differed widely from each other and also from the Zea spp. Presence of greater diversity among the Indian maize collections was also evident from the present analysis.
TL;DR: Pretreatment of the leaves with Triton X-100 at a concentration which specifically inhibits the accessibility of exogenous NAD(P)H to alternative oxidase significantly enhanced the CO response as assessed by in vivo NR assay, supports the hypothesis that the competition for NADH between NR and mitochondrial respiration is regulated by NADH-dehydrogenase located on the outer surface of inner mitochondrial membrane.
Abstract: Differential response in the leaves of tall and dwarf wheat to CO, an inhibitor of cytochrome oxidase and to SHAM, an inhibitor of alternative oxidase appears to be correlated with presence of Rht dwarfing genes This was detected by in vivo nitrate reductase assay after CO treatment and direct O2 uptake in presence of SHAM Pretreatment of the leaves with Triton X-100 at a concentration which specifically inhibits the accessibility of exogenous NAD(P)H to alternative oxidase, Significantly enhanced the CO response as assessed by in vivo NR assay This supports the hypothesis that the competition for NADH between NR and mitochondrial respiration is regulated by NADH-dehydrogenase located on the outer surface of inner mitochondrial membrane
TL;DR: Two RNA binding factors from pea nuclear extracts showed general preferences for single stranded RNA and had a strong preference for poly(U) over the other three homo polymers, suggestive of possible roles for these factors in RNA metabolism in plant nuclei.
Abstract: We have identified and purified two RNA binding factors from pea nuclear extracts. One factor (RBF-1) consisted of at least two polypeptides with molecular weights of approximately 27 and 59 kD; each of these polypeptides could be crosslinked to labelled RNA by ultraviolet light. The other factor (RBF-2) also consisted of two polypeptides (of approximately 93 and 126 kO in size) that could be crosslinked to RNA by UV. Both factors showed general preferences for single stranded RNA. However, RBF-1 displayed a preference for poly(A) and poly(U) over poly(G) or poly(C), while RBF-2 had a strong preference for poly(U) over the other three homo polymers. These properties are suggestive of possible roles for these factors in RNA metabolism in plant nuclei.
TL;DR: For quantitative estimation of coproporphyr in III and protoporphyrin IX from their mixture, a sensitive spectrofluorometric method was developed and it was essential to correct for the fluorescence due to copropoiryrin III at 632 nm and protiporphyrIn IX at 622 nm.
Abstract: For quantitative estimation of coproporphyrin III and protoporphyrin IX from their mixture, a sensitive spectrofluorometric method was developed. At room temperature, coproporphyrin III fluoresces in neutral or alkaline pH at 622 nm having substantial fluorescence at 632 nm where protoporphyrin IX also fluoresces maximally. Similarly, protoporphyrin IX also has substantial fluorescence at 622 nm. Therefore, while estimating protoporphyrin IX (E400 F632) or coproporphyrin III (E400 F622) concentratton, it is essential to correct for the fluorescence due to coproporphyrin III at 632 nm and protoporphyrin IX at 622 nm. This was done by formulating equations from appropriate constants derived from pure samples of coproporphyrin III and protoporphyrin IX. As law as 1 pmole of coproporphyrin III or protoporphyrin IX could be estimated from their mixture by using the spectrofluorometric method.
TL;DR: The complete amino acid sequence of the subunit containing 172 amino acid residues has been established by manual Edman method and the amino acid analyses were carried out.
Abstract: A subunit of molecular weight 18300 has been separated and isolated from seeds of Brassica campestris L. This subunit was cleaved by using cyanogen bromide, trypsin, Staphylococcus aureus V8 protease and chymotrypsin; the fragments obtained from enzymatlc and chemical cleavages were separated and isolated by polyacrylamide gel electrophoresis and gel filtration. The amino acid analyses were carried out. The complete amino acid sequence of the subunit containing 172 amino acid residues has been established by manual Edman method.
TL;DR: A comparable relative efficiency of electron transfer to N2 via nitrogenase under symbiotic as well as ex-planta conditions for both Hup− strain B11 as wellAs Hup+ strain IRBG 46, has been reported and a coordinate regulatory relationship between rubisco and hup genes is suggested.
Abstract: In Azorhizobium caulinodans strain IRBG 46, H2 evolved by nitrogenase Induced uptake hydrogenase ex-planta. The strain expressed an efficient H2 recycling system under both symbiotic and ex-planta conditions. For the first time, a comparable relative efficiency of electron transfer to N2 via nitrogenase under symbiotic as well as ex-planta conditions for both Hup− strain B11 as well as Hup+ strain IRBG 46, has been reported. The study also suggested a coordinate regulatory relationship between rubisco and hup genes.
TL;DR: Two novel cDNAs from Sorghum bicolor have been cloned and sequenced and the sequence analysis revealed that one of the c DNAs, pSbaNS5, is a partial clone which may encode a putative transcription regulator, and its transcript was markedly upregulated by white light.
Abstract: Two novel cDNAs from Sorghum bicolor have been cloned and sequenced. The sequence analysis revealed that one of the cDNAs, pSbaNS5, is a partial clone which may encode a putative transcription regulator, and its transcript was markedly upregulated by white light. The second cDNA, pSbaNS4, may encode a putative glycoprotein of 307 amino acids. Its transcript was present in etiolated tissues and was not significantly altered by white light.
TL;DR: An oxygen sensitive mutant of Azorhizobium caulinodans strain IRBG 46, previously isolated by NTG mutagenesis, has been shown to affect H2- uptake hydrogenase (Hup) activity under symbiotic conditions, but free living Hup activity remained unaffected.
Abstract: An oxygen sensitive mutant of Azorhizobium caulinodans strain IRBG 46 in which N2 fixation ability was affected, was previously isolated by NTG mutagenesis. Now, the mutation has been shown to affect H2- uptake hydrogenase (Hup) activity under symbiotic conditions. However, free living Hup activity remained unaffected. Thus the mutant is Hup- under symbiotic conditions and Hup+ under free living condtions. A possible regulatory link between N2 fixation and H2 uptake system has been discussed.
TL;DR: ‘wick’ method was found to incorporate 5 - and 9- fold higher amount of labelled 35S-methionine into proteins as compared to the other two methods, which was quite effective, safe and easy to use with any kind of tissue or plant species with reproducible results.
Abstract: Three methods for in vivo protein labelling of rice anthers were tested. The ‘wick’ method was found to incorporate 5 - and 9- fold higher amount of labelled 35S-methionine into proteins as compared to the other two methods. This method was quite effective, safe and easy to use with any kind of tissue or plant species with reproducible results. Two dimensional polyacrylamide gel electrophoresis for the qualitative changes in rice anther protein synthesis under water stress was used to test the reproducibility of the method.
TL;DR: The spikelet fertility transformation of this plant was closely related to mean minimum daily and mean average daily temperatures of the 15-d thermosensltive stage prior to 5 days of heading.
Abstract: Unpollinated young ovaries from the F1 plants of the cross between UPRI 95-140, a TGMS donor, and UPRI 95-117, a good plant type line of indica rice were pretreated at 8°C for 14 days and cultured onN6 medium supplemented with lactoprotein hydrolysate 500 mg l-1 2,4-D 4.0 mg l-1 NAA 2.0 mg l-1 and BA 1.0 mg l-1. Five calli with 0.5% of callus induction were obtained. Four of these calli gave complete green plantlets possible of spontaneous dihaploids while the remaining one gave partially green plantlets onMS plant regeneration medium supplemented with casein acid hydrolysate 500 mg l-1, NAA 0.5 mg l-1 and BA 1-5 mg l-1. One of the regenerated green plantlets was identified with thermosensitive genetic male sterility (TGMS) trait. The spikelet fertility transformation of this plant was closely related to mean minimum daily and mean average daily temperatures of the 15-d thermosensltive stage prior to 5 days of heading. It presented complete spikelet sterility between May 1 and Sept 5 and 0.5 to 12.3% and 0.5 to 41.5% of seed set between Sept 18 and Nov 1, 1995 and March 10 to april 27, 1996, respectively at Pantnagar in the northern India. And the line in H2 present homogenous performance with optimum spikelet fertility transformation in Wet season, 1996. This line is being used to develop two-line heterotic hybrids.