ITEMS FROM INDIA

BHABHA ATOMIC RESEARCH CENTRE

Nuclear Agriculture and Biotechnology Division, Mumbai-400085, India.

Combining quality with durable rust resistance and molecular studies in Indian wheat. [p. 48]

B.K. Das, A. Saini, Ruchi Rai, and S.G. Bhagwat (Nuclear Agriculture & Biotechnology Division) and N. Jawali (Molecular Biology Division).

Genetic improvement of wheat for quality and rust resistance is being continued. HMW-glutenin subunits are being used as a criterion for selection. Rust resistance genes such as Sr31 and Sr24/Lr24 are being combined with high-yielding ability. Selections made on the basis of good agronomic characters are being advanced.

A SCAR marker (SCAR30R580bp) for Sr31 gene has been developed. A 1,100-bp band was found associated with the susceptible allele, which also was converted into a SCAR (SCAR 26L1100 bp) marker. The two-marker system is expected to enable identification of homozygous resistant plants in early generation.

Genetic map of bread wheat. [p. 48]

E. Nalini and N. Jawali (Molecular Biology Division), and S.G. Bhagwat (Nuclear Agriculture and Biotechnology Division).

Development of a genetic map of bread wheat based on a population derived from an intervarietal cross is being continued. AFLPs were used to screen the parents and the derived F2. A total of 96 AFLP primer-pair combinations were screened for polymorphism between the parents. All the primer combinations detected polymorphism between the parents with the number of polymorphic bands ranging from 2 to 16. Among these, 14 pair combinations that yielded eight or more number of polymorphic bands in the parents have been used for analyzing the mapping population. In all, 154 polymorphic bands were obtained from 14 primer combinations. These AFLP loci will be integrated in to a map based on STMS, RAPD, and other markers.

Thermotolerance in wheat. [p. 48-49]

Suman Sud, B.K. Das, and S.G. Bhagwat (Nuclear Agriculture and Biotechnology Division).

Heat stress affects performance of wheat plant at early stage and also at the grain-filling stage. Work has been initiated to assess thermotolerance at seedling stage using membrane-stability and cell-viability assays. Over 50 commonly grown cultivars have been tested in both the assays. We found a positive correlation (r = 0.58) between the two assays. Cultivars NIAW-34, PBN-4135-1, and Ajantha were relatively more thermotolerant.

A radiation-induced mutant in wheat. [p. 49]

S.G. Bhagwat (Nuclear Agriculture and Biotechnology Division).

A genetic stock carrying the sphaerococcum character was irradiated with gamma rays, and a lax spike mutant was isolated. The mutant also showed alteration in grain appearance, culm length, and flag leaf blade size (Figure 1). An F2 population from a cross between the two was grown in the field. Flag leaf blade area on main tiller and stomatal frequency on the upper surface were estimated. The entire F2 showed negative correlation as expected. Parental and mutant types were identified on the basis of spike character. These results indicate that the mutant may have stomatal frequency lower than expected as compared to the parent. Because the mutant has alteration in various characters, a large deletion or a mutation with pleiotropic effect may be present.

Publications. [p. 49]

• Bhagwat SG. 2003. Use of image analysis in mutation breeding for changing grain shapes in durum wheat. BARC News Let 237:173-176.
• Nalini E, Jawali N, and Bhagwat SG. 2004. A simple method for isolation of DNA from plants suitable for long term storage and DNA marker analysis. BARC News Let 249:208-214.
• Sharma A, Rao VS, Bhagwat SG, and Bapat MM. 2004. Elongation curve for evaluating chapati making quality of whole wheat flour and its relation with other quality parameters. J Food Sci Tech 41(2):160-164.
• Sud S, Das BK, and Bhagwat SG. 2004. Identification of heat stress tolerant varieties of bread wheat using seedling assays. In: National Symposium on Wheat Improvement for the Tropical Areas, TNAU, Coimbatore, India. 23-25 September, 2004, Book of Abstracts, p. 74.
  

BHARATHIAR UNIVERSITY

Cytogenetics Laboratory, Department of Botany, Coimbatore-641 046, India.

Quality analysis of some Indian hexaploid wheat cultivars. [p. 50]

K. Gajalakshimi and V.R.K. Reddy.

Various physical properties of wheat grain (grain weight, test weight, moisture content, pearling index, particle size index, and grain appearance score), chemical properties of wheat flour (protein, fat, ash, total sugar, damaged starch, and sedimentation value), rheological properties of wheat dough (Farinograph water absorption, dough-development time, stability time, Farinographic resistance, mixing tolerance index, and resistance to extension, extensibility and area of the Extensographic curve), and glutenin quality of seed proteins were evaluated in 50 Indian hexaploid wheat cultivars obtained from various parts of the India in order to assess their suitability for different types of baking and pasta products.

Wheat grains from 44 wheat cultivars showed high to medium physical properties and different chemical constituents. The seeds of these wheat cultivars are semihard to hard in nature, they had higher percentages of protein, total sugar, wet gluten, damaged starch content, and SDS-sedimentation values with lower percentages of fat and ash. These wheat cultivars were recommended for all-purpose use (blending, baking, and pasta making). Grains of the remaining six wheat cultivars showed poor physical properties coupled with lower percentages of protein, total sugar, wet gluten, damaged starch content, and SDS-sedimentation values with higher percentages of fat and ash. These wheat cultivars were recommended for biscuit-making quality.

Rheological properties of wheat dough are recorded using Farinographic and extensographic meters by the Naga Research Institute, Dindigal, Tamil Nadu, India. Wheat dough from 18 wheat cultivars with high dough-development times, stability times, and an area with medium extensibility and low mixing tolerance index values that give high resistance are strong-type wheat doughs. Dough from six wheat cultivars having lower dough-development times, stability times, and area with higher extensibility and mixing tolerance index values are weak-type wheat doughs. Twenty-six wheat cultivars have medium-strong type wheat doughs.

All 50 wheat cultivars also were analyzed for their allelic variations of HMW-glutenin subunit proteins by SDS-PAGE. A total of 11 alleles were identified; three (a, b, and c) at the Glu-A1 locus, five (a, b, c, d, and e) at the Glu-B1 locus, and three (a, b, and d) at the Glu-D1 locus. The most frequent HMW-glutenin subunits were 1 and 2* at Glu-A1, 17+18 at Glu-B1, and 5+10 at Glu-D1. The most frequent protein combinations are 2*, 7+8, 2+12, and 2*, 7, 5+10. The Glu-1 quality score ranged from 5-10. A Glu-1 quality score of 8 is present in large number of the cultivars. We predict that the cultivars that possess high Glu-1 scores, i.e., 9 or above, have the higher glutenin strength needed for blending purposes (mixing with weak quality flour). Cultivars with a Glu-1 score of 8 have good bread-making quality. Glu-1 score below 6 have very good biscuit-making quality. Correlation between the Glu-1 quality score and quality parameters, such as physical, chemical, and rheological parameters, were studied. Significant, positive correlations (p < 0.05) was observed between the Glu-1 score and test weight, sedimentation value, wet gluten, Farinographic parameters, and extensographic parameters (area and resistance) were observed. A negative correlation was found between the Glu-1 score and protein content, mixing tolerance index (degree of softening), extensibility, and the Resistance/Extensibility ratio. Significant positive correlation was found between wet gluten and protein content, sedimentation value, Farinographic parameters and extensographic parameters (except MTI). Significant negative correlation was found between wet gluten and test weight and mixing tolerance index.

Based on the results of physical properties of the grain, chemical properties of flour, rheological properties of dough, and glutenin-subunit composition of seed proteins in the 50 Indian hexaploid wheats, the cultivars were ordered into five groups: high, medium-high, medium, medium-low, and low quality wheats. Among the 50 wheat cultivars, 13 were high quality, five were medium-high, 22 were medium, 4 were medium-low, and 6 were low quality.

Publication. [p. 51]

• Kalaiselvi G and Reddy VRK. 2005. Yield performance of the near-isogenic lines carrying different rust resistance genes in three Indian wheat cultivars. Res Crops 6(1):In press.
  

CH. CHARAN SINGH UNIVERSITY

Department of Agricultural Botany, Meerut - 250 004, India.

P.K. Gupta, H.S. Balyan, R. Bandopadhyay, N. Kumar, S. Sharma, P.L. Kulwal, S. Rustgi, R. Singh, A. Goyal, and A. Kumar.

Development and use of molecular markers for wheat genomics. [p. 51-54]

QTL analysis for different traits using trait-specific, intervarietal mapping populations.
QTL analysis for preharvest sprouting tolerance (PHST) using trait-specific, intervarietal mapping populations.
QTL analysis for PHST in bread wheat was earlier conducted by us following single locus composite interval mapping (CIM) and two locus analysis (QTLMapper), using an International Triticeae Mapping Initiative population (ITMIpop). In this study, an intervarietal mapping population in the form of RILs developed from a cross between the genotypes, SPR8198 (PHS tolerant) and HD2329 (PHS susceptible) was used for single-locus CIM. The parents and the RIL population were grown in six different environments, and the data on PHS were recorded on a scale of 1-9 with a score of 1 for genotypes with complete resistance to PHST and a score of 9 for the genotypes with complete sprouting in each case. A framework linkage map of chromosome 3A with 13 markers was prepared and used for QTL analysis. A major QTL (QPhs.ccsu-3A.1) was identified on 3AL explaining 24.68 % to 35.21 % of the variation in an individual environment (for details, see Table 1). When PHST data from six environments was pooled, the QTL explained 78.03 % variation. The results obtained here are significant, because the QTL detected seems to be new and was present in all the environments and also with the pooled data, a rather rare event in QTL analysis. The positive additive effects in the present study suggest that a superior allele of the QTL is available in the superior parent (SPR8198), which can be used for MAS for the transfer of this QTL allele to elite strains with to obtain superior progeny. This work has been submitted for publication in Plant Science, and is currently under review.

QTL analysis for yield and its component traits, using a trait-specific, intervarietal mapping population for grain weight (GW).
QTL analysis for yield and its four component traits (tillers per plant, grain yield, grains/spike, and GW) in bread wheat was conducted following CIM and using an intervarietal mapping population for grain weight in the form of RILs developed from a cross between Rye Selection 111 (high GW) and Chinese Spring (low GW). The parents and the RIL population were grown in six different environments, and the data on the four traits were recorded. For QTL interval mapping, framework linkage maps were prepared for chromosomes 1A, 2A, 2B, and 7A using 68 markers, including SSRs, AFLP, and SAMPL. For four different yield and component traits, the number of QTL ranged from three for grain weight to five each for tillers/plant and grain yield. A total of 17 QTL were identified. Of 17 QTL, only five were considered as consistent QTL, because they were detected in more than three environments in this study (for details, see Table 1). Of these five QTL, three for GW were detected, one each on 1A, 2B, and 7A, whereas one each for grain yield and grains/spike was detected, both on chromosome 2B. No consistent QTL was detected for tillers/plant.

QTL analysis for agronomically important traits using a trait specific intervarietal mapping population for grain protein content (GPC).
QTL analysis for growth related traits. QTL interval mapping for four growth traits (days-to-heading, days-to-maturity, early growth habit, and plant height) were made using an intervarietal mapping population for GPC. CIM was done, using QTL Cartographer, for all the above four growth traits with phenotypic data scored in six different environments. Our earlier studies on QTL analysis for these traits were confined to a single environment and involved an ITMI population (Kulwal et al. 2003). The number of QTL detected ranged from seven for plant height to 21 for days-to-heading. A total of 64 QTL for all of the four traits were detected (for details, see Table 1). QTL that were consistent across the environments included one each for days-to-heading and days-to-maturity, both located on chromosome 5B. No consistent QTL were detected for early growth habit or plant height.

QTL interval mapping for yield and yield-contributing traits. For eight different yield and yield-contributing traits, QTL interval mapping (CIM) was by QTL Cartographer using data collected on RILs (GPC population) grown in six different environments. For different traits, the number of QTL ranged from seven, for grain weight, to 17, for grain yield, with a total of 92 QTL for all the eight traits (for details, see Table 1). One consistent QTL each were detected for spike length on chromosome 2B and for grain weight on chromosome 3A. No consistent QTL were detected for the remainder of the traits.

High-resolution mapping of the genomic regions containing an important QTL for GPC.
For the purpose of high-density mapping of an important QTL for grain-protein content, an F2 population of about 2,000 plants was derived from a cross between two RILs, one of them containing high GPC alleles and the other containing low GPC alleles for the selected QTL. DNA was isolated from individual F2 plants, which are being genotyped with the markers flanking the selected QTL to identify recombinants for the targeted region. So far, 19 F2 homozygous recombinants for the markers flanking the selected QTL have been identified.

Developing and using EST-SNPs and anchored SSRs in bread wheat.
Developing, validating, and using EST-SNPs.
As is widely known, more than 580,000 ESTs are now available for bread wheat. This resource was used for the development of SNPs under the umbrella of the Wheat SNP Consortium (WSC). Forty-eight EST-contigs, each having 20 to 89 ESTs, were searched for the presence of HSVs (homoeologue specific variations) and SNPs. In this study, 462 HSVs were detected in 47 EST-contigs, allowing subclustering of the 47 EST-contigs into 174 subcontigs and facilitating detection of 230 putative SNPs in 42 EST-contigs. An average density of one SNP every 273.9 bp was calculated. Out of 230 SNPs, 123 (53.5 %) represented transitions, and the remaining 107 (46.5 %) represented transversions, suggesting that transitions are relatively more frequent than transversions. In this study, 42 locus specific STS primers were designed and used for PCR with genomic DNA from 30 diverse bread wheat genotypes. Only 39 (92.8%) primers amplified fragments in 15 to 30 genotypes. The remaining three primers failed to give any product. Ten of the 39 primers each amplified a solitary band (representing a single homoeolocus) in each of 237 (79 %) of the 300 possible primer-genotype combinations (10 primers x 30 genotypes). Only the above 10 primers were considered suitable for validation of 30 putative SNPs that were detected in silico in the amplifiable region (amplicon) of the corresponding EST-contigs. Out of 30 putative SNPs, only seven SNPs were validated; however, eight new SNPs also were detected through direct sequencing of PCR products from 30 genotypes.

The above 15 SNPs (seven validated + eight new) detected in this study allowed construction of 11 haplotypes. The above data also was used for the construction of a dendrogram to study genetic similarity/diversity.

Developing and using anchored SSRs.
A large number of SSR markers (wmc markers) were earlier developed under the aegis of an international effort 'Wheat Microsatellite Consortium (WMC)'. In this exercise, although the sequences of as many as ~1,200 clones were found to contain SSRs, primers could be designed for only ~600 SSRs, leaving another ~600 sequences that either had poor quality or were considered unsuitable for designing of primers mainly due to the occurrence of SSRs too close to an end of the sequence. We utilized a part of these sequences for designing 52-anchored SSR primers. In this study, a set of 105 52-anchored SSRs were developed. These 105 52-anchored SSR primers were used for developing STMS markers. A subset of 45 of these anchored SSR primers also was used for microsatellite-anchored fragment length polymorphism (MFLP) analyses. In the STMS analysis, the proportion of functional anchored-SSR primers was close to that reported earlier for simple SSR primers and in MFLP analysis. The average number of polymorphic bands per primer combination was 11.9, although anchored-CT/GA SSR primers gave a relatively higher average number of polymorphic bands (17.88). The above MFLP data also was used for genetic diversity analysis among eight bread wheat genotypes (representing parents of four intervarietal mapping populations available from our laboratory). The average polymorphic information content (PIC) was found to be 0.057, and the average genetic similarity (GS) was 0.451.

Use of C0t fractionation (CF) and methyl filtration (MF) for genomics research in bread wheat.
To demonstrate the utility of C0t fractionation (CF) and methyl filtration (MF) in assaying wheat genome complexity, a rather small fraction (671.67 kb) was sequenced and analyzed for the presence of protein-coding genes, noncoding (nc) RNA genes, SSRs, and transposable elements (TEs). Results demonstrated the utility of gene-enrichment techniques (high C0t and MF) in assaying comparatively large number (12-fold) of genes. The above gene-enrichment techniques still retain as much as more than 23 % repeat elements. Of most interest are the fractions assayed by CF and MF, which vary substantially for repeat and low-copy content, indicating the need of using both of the above techniques in parallel to study entire genome complexity of wheat.

The SSRs were three times more abundant in low-copy (high C0t and MF) fractions of wheat genome than in the repeat (reassociated DNA; RD) fraction. We also observed that low-copy sequences have more trinucleotide SSRs, particularly those with the 'CCG/CGG' motif. These results conform with our earlier results on wheat EST-SSRs (Gupta et al. 2003).

A large proportion of MF, high C0t and RD sequences also were similar with already known miRNA (micro RNA) sequences available in the public domain. Target mRNAs for a large proportion of the candidate miRNAs also could be detected on the basis of their similarity with wheat ESTs, further confirming their validity and presence in low-copy, transcriptionally active, hypomethylated regions of the genome.

Physical mapping of SSRs on all the 21 chromosomes of bread wheat.
Approximately 2,150 SSRs loci already have been mapped genetically in bread wheat, but only ~1,050 SSR loci have been mapped physically leaving more than ~1,100 SSR loci that have not been placed on the physical maps so far. In the present study, 42 nullisomic-tetrasomic, 24 ditelocentric, and 164 homozygous overlapping deletion lines from a total of 436 available deletion lines were used for bin localization of SSRs. A set of 527 SSRs was tried, leading to successful mapping of 270 SSRs on 313 loci covering all the 21 chromosomes. A maximum of 119 loci (38 %) were located in the B genome, and a minimum of 90 loci (29 %) mapped in the D genome. Similarly, homoeologous group 7 had a maximum of 61 loci (19 %), and group 4 a minimum of 22 loci (7 %). Of the 270 SSRs, 39 SSRs had multiple loci, but only eight of these detected homoeologous loci. The linear order of loci in physical maps largely corresponded to those on the genetic maps. Apparently, distances between each of only 26 pairs of loci significantly differed from the corresponding distances on genetic maps. Some loci, which were genetically mapped close to the centromere, were physically located distally, whereas other loci that were mapped distally in the genetic maps were located in the proximal bins in the physical maps. This result suggested that although the linear order of the loci was largely conserved, variation does exist between genetic and physical distances.

Radiation hybrid (RH) mapping in bread wheat.
In order to determine the linear order of markers placed in a particular bin, more deletions were induced by irradiating seeds of monosomic lines for chromosomes 1A, 2A, and 3A of bread wheat cultivar Chinese Spring. Seeds of the above monosomic lines were exposed to three different dosages of gamma rays (30, 40, and 50 krad) to find out optimum dosage required for induction of maximum number of chromosome breakages at minimum mortality rate. Irradiated seeds were sown in the field, and DNA was isolated from first four tillers of each plant. PCR analysis of the irradiated monosomic plants is being conducted using 10, 16, and 19 SSR markers already physically mapped on bread wheat chromosome 1A, 2A, and 3A, respectively (Goyal et al. 2005). This study will help in further subdividing the available deletion bins, to help further fine physical mapping.

Future work.
Marker-assisted selection for high GPC and PHST.
We are in the process of introgression of one QTL each for high GPC and PHST into elite, Indian bread wheat genotypes with low GPC and susceptibility to PHS through backcrossing programs. In the backcrossing program, we are using Yecora Rojo as donor parent for high GPC (procured from J. Dubcovsky of University of California, Davis, USA) and SPR8198 as donor parent for PHST (procured from Punjab Agric Univ, Ludhiana, India). The F1 plants derived from the crosses between several recipient patents (K9107, HD2687, Raj3765, PBW373, HI977, HD2329+Lr24+Lr28, PBW343+Lr9, PBW343+Lr19, and PBW343+Lr24) with the above two donor parents are already raised in field and backcrosses will be done in this season.

High-density mapping.
High-density mapping of 3.4 cM on 2D carrying a major QTL for high GPC. The F2 plants that were homozygous recombinants for the markers flanking the selected QTL will be further used for high-density mapping of the selected region. SAMPL, AFLP, STS, and SSR markers will be used for genotyping of the selected homozygous recombinant F2 plants to saturate the selected region. Wheat EST markers already mapped in the targeted region and those mapped in the syntenous regions in other grasses also will be used for saturation mapping of the selected region. More SSRs will be developed from an arm-specific library and will be used for saturation mapping of the targeted region.

High-density mapping of 17 cM on 3A carrying a major QTL for PHST. Two strategies will be followed for high-density mapping of a 3A region carrying a major QTL for PHST. In the first approach, we will develop different EST-derived markers from the ESTs already physically mapped in the bin (3AL-3) to which our QTL belongs. We also are trying to ascertain the putative order of the above EST-derived markers using sequence of rice chromosome 1 as a reference. In the second approach, we are developing molecular markers from the chromosome arm (3AL) specific library developed in a collaborative project with J. Dolezel, Olomouc, Czech Republic. The above exercises will lead to enrichment of the interval containing QTL of interest with molecular markers, which may ultimately lead to the isolation of the QTL of interest.

References.

• Kulwal PL, Roy JK, Balyan HS, and Gupta PK. 2003. QTL mapping for growth and leaf characters in bread wheat. Plant Sci 164:267-277.
• Gupta PK, Rustgi S, Sharma S, Singh R, Kumar NK, and Balyan HS. 2003. Transferable EST-SSR markers for the study of polymorphism and genetic diversity in bread wheat. Mol Genet Genomics 270:315-323.
  

Publications. [p. 54-55]

• Bandopadhyay R, Sharma S, Rustgi S, Singh R, Kumar A, Balyan HS, and Gupta PK. 2004. DNA polymorphism among 18 species of Triticum-Aegilops complex using wheat EST-SSRs. Plant Sci 166:349-356.
• Goyal A, Bandopadhyay R, Sourdille P, Endo TR, Balyan HS, and Gupta PK. 2005. Physical mapping of microsatellite markers (SSRs) in bread wheat genome with the help of deletion stocks and its relationship with genetic maps. Funct Integr Genomics (Submitted).
• Gupta PK and Dhar MK. 2004. Advances in molecular cytogenetics: potential for crop improvement. In: Plant Breeding - Mendelian to Molecular Approaches (Jain HK and Kharakwal MC, Eds). Narosa Publishing House, New Delhi. Pp. 97-114.
• Gupta PK and Rustgi S. 2004. Molecular markers from the expressed region of the genome in higher plants. Funct Integr Genomics 4:139-162.
• Gupta PK and Varshney RK. 2004. Cereal Genomics. Kluwer Scientific Publishers, The Netherlands. 639 pp.
• Gupta PK and Varshney RK. 2004. Cereal genomics: An overview. In: Cereal Genomics (Gupta PK and Varshney RK, Eds). Kluwer Academic Publishers. Pp. 1-18.
• Gupta PK, Balyan HS, Bandopadhyay R, Kumar N, Sharma S, Kulwal PL, Rustgi S, Singh R, Goyal A, and Kumar A. 2004. Development and use of molecular markers for wheat genomics. Ann Wheat Newslet 50:52-55.
• Gupta PK, Kulwal PL, and Rustgi S. 2005. Wheat cytogenetics in the genomics era and its relevance to breeding. In: Plant Cytogenetics (Puertas MJ and Naranjo T, Eds). Cytogenet Genome Res (In press).
• Gupta PK, Rustgi S, and Kulwal PL. 2005. Linkage disequilibrium and association studies in higher plants: Present status and future prospects. Plant Mol Biol (In press).
• Gupta PK, Sharma S, Kumar S, Balyan HS, Beharav A, and Nevo E. 2004. Adaptive ribosomal DNA polymorphism in wild barley at a mosaic microsite, Newe Ya'ar in Israel. Plant Sci 166:1555-1563.
• Gupta PK. 2004. Aneuploid mapping in diploids. In: Encyclopedia of Plant and Crop Science (Goodman RM, Ed), Marcel Dekker Inc. Pp. 33-36.
• Gupta PK. 2004. Molecular Biology and Genetic Engineering. Rastogi Pub, Meerut, India. 590 pp.
• Gupta PK. 2004. Molecular cytogenetics of wheat. In: Proc 2nd Internat Group Meet, Wheat Technologies for Warmer Areas (Rao VS, Singh G and Misra SC, Eds). Anamya Publishers, New Delhi. Pp. 124-137.
• Gupta PK. 2004. The 2004 Nobel Prize in Physiology or Medicine: Odorant receptors and organization of the olfactory system. Curr Sci 87:1500-1504.
• Kole C and Gupta PK. 2004. Genome mapping and map based cloning. In: Plant Breeding-Mendelian to Molecular Approaches (Jain HK and Kharakwal MC, Eds). Narosa Publishing House, New Delhi. Pp. 257-300.
• Kulshreshtha R, Kumar N, Balyan HS, Gupta PK, Khurana P, Tyagi AK, and Khurana JP. 2005. Structural characterization, expression analysis and evolution of the red/far-red sensing photoreceptor gene, PHYTOCHROME C (PHYC), localized on the 'B' genome of hexaploid wheat (Triticum aestivum). Planta (In press).
• Kulwal P, Kumar N, Gaur A, Khurana P, Khurana JP, Tyagi AK, Balyan HS, and Gupta PK. 2005. Mapping of a major QTL for pre-harvest sprouting tolerance on chromosome 3A in bread wheat. Plant Sci (Submitted).
• Kulwal PL, Kumar N, Kumar A, Gupta RK, Balyan HS, and Gupta PK. 2005. Gene networks in hexaploid wheat: interacting quantitative trait loci for grain protein content. Funct Integr Genomics DOI 10.1007/s10142-005-0136-3.
• Kulwal PL, Singh R, Balyan HS, and Gupta PK. 2004. Genetic basis of pre-harvest sprouting tolerance using single-locus and two-locus QTL analyses in bread wheat. Funct Integr Genomics 4:94-101.
• Kumar N, Balyan HS, and Gupta PK. 2005. Early testing of intervarietal chromosome substitutions for grain protein content in bread wheat and its utility in QTL analysis. Plant Breed (Submitted).
• Roy JK, Lakshmikumaran MS, Balyan HS, and Gupta PK. 2004. AFLP-based genetic diversity and its comparison with diversity based on SSRs, SAMPL and phenotypic traits in bread wheat. Biochem Genet 42:43-59.
• Schulman A, Gupta PK, and Varshney RK. 2004. Organization of retrotransposons and microsatellites in cereal genomes. In: Cereal Genomics (Gupta PK and Varshney RK, Eds). Kluwer Academic Publishers, Dordrecht, the Netherlands. Pp. 83-118.
• Sharma S, Beharav A, Balyan HS, Nevo E, and Gupta PK. 2004. Ribosomal DNA polymorphism and its association with geographical and climatic variables in 27 wild barley populations from Jordan. Plant Sci 166:467-477.
• Sharma S, Kumar S, Balyan HS, and Gupta PK. 2004. Interlocus homogenisations of ribosomal DNA repeat units in barley. Curr Sci 86:384-386.
• Singh R, Rustgi S, Bandopadhyay R, Kumar N, Sharma S, Balyan HS, and Gupta PK. 2005. Anchored-SSRs for the study of DNA polymorphism and genetic diversity in bread wheat. Plant Mol Biol Reptr (Submitted).
• Varshney RK, Balyan HS, and Langridge P. 2005. Molecular markers and maps for wheat genetics, breeding and genomics. In: Genome mapping and molecular breeding I. Cereals (Kole C, Ed). Oxford & IBH Publishers, New Delhi (In press).
  

 

 

CHAUDHARY CHARAN SINGH HARYANA AGRICULTURAL UNIVERSITY

Department of Plant Pathology, Hisar-125004, India.

 

Effect of sowing date on the interaction of loose smut and flag smut of Indian wheat cultivars. [p. 56-57]

Rajender Singh, M.S. Beniwal, and S.S. Karwasra.

Introduction. Simultaneous occurrence of loose smut and flag smut of wheat has been reported (Aujla and Sharma 1997; Bedi et al. 1959). These authors reported that flag smut-infected plants showed twisting and bending of coleoptile in the seedling stage with the formation of bleached spots on the coleoptile. Some plants produce smutted heads that emerged later than healthy spikes. All the spikes on the affected plants were smutted and produced very few tillers. Some plants were infected with both smuts. No information is available in the literature on the interaction of smuts on different wheat cultivars grown over India. The present investigation was made under field conditions.

Materials and Methods. A field experiment was conducted at Plant Pathology research area of Plant Pathology research of area of CCSHAU-Hisar during 1997-2000 crop season. Loose smut spores were artificial inoculated in previous crop season in all 25 cultivars so as to serve as loose smut infected seed for next crop season. Each subplot for a cultivar had six rows of 2-m length. Sowing dates of 25 November and 15 December were used for all treatments. Loose smut inoculated seeds were smeared with flag smut teliospores at 2 gm/100 gm of seed having 78 % viability. All normal agronomic practices were followed. Disease incidence was observed as the percent infected tillers in each subplot. Tillers having both diseases were counted for both diseases separately.

Results and Discussion. Results of this study are found in Table 1. Ustilago segetum var. tritici is internally seedborne, but Urocystis agropyri is an external soil contaminant. Both pathogens move side by side systemically in the same plant. The highest loose smut incidence (46.66 %) was found on Sonalika, followed by Raj 3765, but Sonalika showed resistance to flag smut. The highest flag smut incidence was observed on UP2338, followed by PBW 435 (16.66 %) under normal sowing conditions. A delay in sowing drastically reduced the incidence of both diseases. Sonalika had the highest incidence of loose smut (27.57 %) followed by HD 2687 (26.16 %). The maximum incidence of flag smut was in UP 2338, followed by WH 416. Two cultivars, WH 896 and Raj 1555, had resistance against loose smut and flag smut, whereas Sonalika, WH 283, WH291, and HD2329 also exhibited resistance to flag smut. Interestingly, the same tiller expressed both diseases separately. Loose smut incidence was predominant over flag smut in same cultivar, even though flag smut symptom appeared before those of loose smut. Delayed sowing caused less disease incidence, because of fungus inactivation or less spore germination accompanied with falling temperatures. The same observations were made by Beniwal et al. (1992) and Bedi et al. (1959), confirming our results.

Table 1. The effect of sowing date on the interaction of loose smut and flag smut of wheat at three different sowing dates, CCS Haryana Agricultural University, Hisar.

Cultivar	Percent disease incidence
	25 November		5 December		30 December
	Loose smut	Flag smut	Loose smut	Flag smut	Loose smut	Flag smut
C306	33.57	12.50	26.33	11.88	21.44	9.83
Sonalika	46.66	1.71	34.22	0.90	27.57	0.50
WH 147	36.55	16.35	28.44	14.83	22.75	11.83
WH 157	28.44	12.50	25.83	11.66	18.66	9.16
WH 283	25.33	0.00	16.66	0.00	10.33	0.00
WH 291	28.77	0.00	21.33	0.00	18.33	0.00
WH 416	33.33	18.11	28.33	14.33	19.83	12.11
WH 533	26.11	13.33	21.66	10.75	19.66	8.33
WH 542	29.66	16.33	26.66	12.33	23.33	9.33
WH 896	0.00	0.00	0.00	0.00	0.00	0.00
Sonak	26.66	11.66	21.33	10.66	16.66	9.66
HD 2009	24.33	14.33	18.66	11.88	12.33	10.16
HD 2285	24.66	14.66	16.66	12.33	12.83	10.83
HD 2329	27.93	2.83	18.66	0.00	13.33	0.00
HD 2687	34.28	14.57	29.83	14.71	26.16	11.57
PBW 175	23.66	16.33	20.77	12.83	19.14	10.11
PBW 343	35.44	12.50	29.16	10.66	18.11	8.11
PBW 373	37.77	16.25	31.57	13.66	26.33	10.33
PBW 435	38.33	16.66	31.44	12.83	24.33	10.66
Raj 1555	0.00	0.00	0.00	0.00	0.00	0.00
Raj 3077	37.33	11.33	28.33	9.83	22.67	7.66
Raj 3765	38.66	10.33	29.66	9.11	21.77	8.96
Raj 3777	33.66	10.57	26.33	8.83	21.44	7.83
UP 2338	29.66	19.83	17.83	16.33	14.57	13.44
UP 2425	26.77	12.57	21.66	10.11	16.57	8.57
C.D. (0.5 %)	2.98	3.45	3.78	2.87	3.11	2.67

References.

• Aujala SS and Sharma YR. 1977. Simultaeous occurrence of Ustilago nuda tritici and Urocystis agropyri. Indian Phytopath 30:262.
• Bedi KS, Chohan JS, and Chahal DS. 1959. Simultaneous occurrence of Ustilago tritici (Pers) and Anguina tritici in a single ear of wheat. Indian Phytopath 12:187.
• Beniwal MS, Karwasra SS, and Parashar RD. 1992. Effect of sowing date on the incidence flag smut of wheat in Haryana. Crop Res 5(3):598-600.

Effect of sowing date on the interaction of flag smut and earcockle of wheat cultivars. [p. 57]

Rajender Singh, M.S. Beniwal, and S.S. Karwasra.

The simultaneous occurrence of earcockle (Anguina tritici ) with the fungus Urocystis agropyri was reported by Bedi et al. (1959), Aujla and Sharma (1977), and Pruthi and Bhatti (1982). Whether or not concomitant occurrence of the fungus with nematode has any synergistic or antagonistic effect on the development of disease incidence is unknown. Therefore, we attempted to study effect of sowing on concomitant of flag smut and earcockle on wheats cultivated in India.

A field experiment was conducted at Plant Pathology research farm of CCSHAU-Hisar during the 1997-2000 crop season. Before sowing, seed of each cultivar were smeared with dry teliospores powder having 78 % viability at 2-gm inoculum/100 gm of seed. Each subplot for a cultivar had six 2-m rows. Sowing dates of 25 November, 15 December, and 30 December were used for all treatments. Each row received 10 nematode galls. All normal agronomic package practices were followed. Disease incidence was observed as a percent infected tiller basis in each subplot. The same tiller having both diseases were counted separately.

The highest flag smut incidence (15.37 %) was observed on Raj3765, followed Raj 3777 (14.96 %) at normal sowing, but in delay sowing, disease incidence declined to 10.87 and 9.91%, respectively (Table 2). WH283, WH291, WH896, HD2329, and Raj1555 were found to be resistant in normal and late sowing, possibly because of falling temperatures that prolonged the time of teliospore germination. Conversely, with delayed sowing (30 December), earcockle incidence increased to 38.83 % (HD 2285) but decreased at the normal sowing date (25 November). Anguina tritici have more time for infection with a prolonged time for seed germination. If both flag smut and earcockle were observed on same tiller, they were counted separately. In normal and late sowings, earcockle incidence predominated over flag smut. The same observations were made by Pruthi and Gupta (1986) and Beniwal et al. (1992) and confirm our results. Pruthi and Gupta (1986) reported that the presence of fungi with nematodes has an adverse effect on the number, motility, and development of larvae at normal sowing time.

References.

• Aujla SS and Sharma YR. 1977. Simultaneous occurrence of Ustilago nuda tritici and Urocystis agropyri. Indian Phytopath 30:262.
• Bedi KS, Chohan JS, and Chahal DS. 1959. Simultaneous occurrence of Ustilago tritici and Anguina tritci in a single ear of wheat. Indian Phytopath 12:187.
• Pruthi IJ and Gupta DC. 1986. Studies on combined occurrence of Ustilago tritici and Anguina tritici. Haryana Agric Univ J Res SVI(4):392-394.

Effect of sowing date on the interaction of loose smut, flag smut, and earcockle of wheat cultivars grown in India. [p. 58-60]

Rajender Singh, M.S. Beniwal, S.S. Karwasra, and Sher Singh.

Introduction. The simultaneous occurrence of loose smut, flag smut, and earcockle of wheat has been reported (Bedi et al. (1959) and Aujla and Sharma (1977), who reported that flag smut infected plants/tillers showed twisting and bending of the coleoptile in the seedling stage with formation of bleached spots on the coleoptile. The same plant produced smutted spikes that emerged later than the healthy ones. Plants were infected with both smuts. They also noticed that two-thirds of the lower spikes were nearly totally filled with black loose smut sori. The upper portion of this spike was infected by earcockle, the black galls of which were clearly visible. No information is available in the literature on the interaction of smuts and earcockle of the different wheats cultivated in India and the effect of different sowing dates. The present investigation was made under field condition.

Materials and Methods. We conducted a field experiment at the Plant Pathology Research Area of CCSHAU-HISAR during 1997-2000 Loose smut spores were artificially inoculated in the previous crop season in all cultivars to serve as inoculum for next crop season. Each cultivar subplot was 6 2-m rows. Sowing dates, 25 November, 15 December, and 30 December, were used for all treatments. Loose smut-inoculated seed was inoculated with teliospores at 2 gm/100 gm seed and sown. Each cultivar had 78 % viability in each subplot. Each row also received 20 nematode galls. All normal growing practices were followed. Disease incidence was observed on a percent infected tiller basis in each subplot. If the same tiller had symptoms for all diseases, each was counted separately.

Results and Discussion. Loose smut is internally seed borne, but Urocystis agropyri and Anguina tritici are externally seed borne. All inoculula move systemically in parallel in same plant. Loose smut-infected spikes emerged earlier than healthy spikes. Flag smut symptoms appeared first 45 days-after-sowing, and were followed by earcockle and loose smut symptoms (Table 2 and Table 3). At the normal sowing time, the highest incidence of loose smut was observed in Sonalika (43.18 %) followed by C-306 (39.17 %) but was reduced to 32.5 % and 28.5 %, respectively, at later sowings. No loose smut or flag smut was found on WH 896 and Raj 1555. Sonalika, HD 2329, WH 283, WH 291, WH 896, and Raj 1555 also expressed resistance against flag smut. The highest flag smut (28.18 %) and earcockle incidence (14.57 %) was observed in HD 2285 in a normal sowing but with a delay in sowing, flag smut incidence declined to 9.14 % and earcockle incidence increased to 36.66%. Disease incidence in Raj 3765 (9.14 %) also declined (8.44) at later sowing dates. Sonalika, HD 2329, WH 283, WH 291, WH 896, and Raj1555 expressed resistance against flag smut in all sowing times. None of the cultivars was resistant to earcockle. HD 2285 had the maximum earcockle incidence (20.14 %) followed by Raj 1555 (20.11 %), when sown late. However, loose smut incidence was predominant over earcockle and flag smut. We noticed that with later sowings, loose smut and flag smut incidence was adversely affected, whereas earcockle incidence was enhanced. A reduction in teliospore germination along with falling temperatures in November and December and the presence of the nematode A. tritici may provide more time for infection due to a prolonged germination time. Similar observations were made by Beniwal et al. (1992) and Pruthi and Gupta (1986) on single spikes, which confirm our results. Pruthi and Gupta (1986) reported that the presence of fungus and nematodes has an adverse affect on the number, motility, and development of larvae. In some plants, the same tiller showed symptoms of all three diseases.

Table 2. The effect of sowing date on the interaction of flag smut and earcockle of wheat cultivars, CCS Haryana Agric Univ, Hisar.

Cultivar	Percent disease incidence
	25 November		5 December		30 December
	Flag smut	Earcockle	Flag smut	Earcockle	Flag smut	Earcockle
C306	12.81	2.33	11.11	26.14	7.58	31.22
Sonalika	2.57	16.46	1.83	21.16	1.16	28.27
WH 147	14.96	20.94	10.86	28.76	8.45	35.53
WH 157	12.93	17.88	11.93	20.44	9.57	25.44
WH 283	0.00	20.81	0.00	28.33	0.00	35.11
WH 291	0.00	21.11	0.00	29.14	0.00	37.44
WH 416	13.47	17.07	11.61	21.33	8.36	26.17
WH 533	12.11	18.77	10.81	21.57	8.84	28.25
WH 542	14.66	17.11	11.27	21.27	9.61	25.33
WH 896	0.00	10.16	0.00	14.44	0.00	16.66
Sonak	11.33	16.44	10.43	21.57	8.36	27.11
HD 2009	13.24	18.94	11.91	21.71	8.75	28.14
HD 2285	16.66	21.44	13.83	29.81	10.93	38.83
HD 2329	2.83	20.66	1.93	3.66	1.33	30.16
HD 2687	11.16	19.57	10.66	22.83	8.93	29.57
PBW 175	10.25	17.11	6.83	20.83	5.55	26.44
PBW 343	14.77	18.33	11.44	21.77	9.11	27.53
PBW 373	10.81	15.77	9.91	18.16	8.16	21.11
PBW 435	11.37	18.93	8.73	22.33	6.93	24.77
Raj 1555	0.00	13.57	0.00	16.73	0.00	18.87
Raj 3077	14.86	20.33	11.28	24.97	9.88	27.83
Raj 3765	15.37	20.34	11.71	25.66	10.87	28.77
Raj 3777	14.96	19.93	12.93	24.33	9.91	29.33
UP 2338	12.88	18.33	10.11	21.88	8.73	27.28
UP 2425	14.16	17.44	11.25	20.93	8.77	26.84
C.D. (0.5 %)	2.56	3.11	3.67	2.98	3.89	3.96

References.

• Aujala SS.and Sharma YR. 1977. Simultaneous occurrence of Ustilago nuda tritici and Urocystis agropyri. Indian Phytopath 30:262.
• Bedi KS, Chohan JS, and Chahal DS. 1959. Simultaneous occurrence of Ustilago tritici (Pers) and Anguina tritici in a single ear of wheat. Indian Phytopath 12:187.
• Beniwal MS, Karwasra SS, and Parashar RD. 1992. Effect of sowing date on the incidence of flag smut of wheat in Haryana. Crop Res 5(3):598-600.
• Pruthi IJ and Gupta DC. 1986. Studies on combined occurrence of Ustilago tritici and Anguina tritici. Haryana Agric Univ J Res XVI 4(3):392-394.

Influence of plant extracts on teliospore germination of Neovossia indica in wheat. [p. 60-61]

Rajender Singh, S.S. Karwasra, and M.S. Beniwal.

Of eight plant extracts tested, the maximum inhibition of teliospore germination was found in neem (Azadirachta indica), up to 91.75 %, followed by onion (83.35 %). The minimum level of inhibition was observed in Cannabis sativa (38.35 %). Plant extracts are known to have antimicrobial properties and are easily available, ecofriendly, and cheap. The use of extracts from higher plants for controlling diseases dates to 470 BS (Sharvelle 1963). The Karnal bunt pathogen perpetuates in the seed and soil in the form of teliospores. No chemical is able to completely inhibit teliospore germination. Water extracts from plants were evaluated for their effect on teliospore germination.

Materials and Methods. Plant extracts were prepared by macerating leaves or bulbs in distilled water at 1:1 (w/v) from ak (Calotropis procera, leaves), datura (Datura metel, leaves), safeda (Eucalyptus globules, leaves), onion (Allium cepa, leaves), bhang (Cannabis sativus. leaves), neem (Azadirachta indica, leaves), garlic (Allium longicuspus, bulb), and zinger (Zingiber officinale, leaves). After dilution of the standard extract to 1 %, 0.5 %, and 0.25 %, teliospore germination was recorded. In one treatment, the teliospores from bunted seed were directly sprinkled over the different concentration of the plant extract solutions to evaluate their germination percentage. In another experiment, teliospores from treated seeds were sprinkled over distilled water in the petriplates and incubated at 20 C for 20 days. Teliospore germination was recorded weekly by recording the total and the number of germinated teliospores scored under a microscopic field (10 x 10). The percent teliospore germination was calculated. For all dilutions, bunted seeds were dipped for 48 hrs to evaluate their efficacy against seed-borne inoculum lying protected beneath the pericarp. Simultaneously, seed germination also was tested.

Results and Discussion. Although there was no adverse effect of the plant extracts on seed germination, the treatments were ineffective at eliminating seed-borne inoculum (Table 4) from broken seed dipped in neem, onion, and garlic extracts. Teliospores from within the intact pericarp of the treated seeds germinated. The lowest teliospore germination was observed in extracts from neem leaves, but none of plant extracts completely inhibited teliospore germination. Extracts from garlic, onion, and neem were effective in making the teliospores unviable, but it could not penetrate the pericarp of the seed. Bulb extracts of garlic have been found inhibitory to T. indica and R. solani (Sundaraj et al. 1998; Sharma and Nanda 2000). Our investigation may vary due to different isolates found at Hisar.

References.

• Sharvelle EC. 1963. The nature and use of modern fungicides. Burgess Publishing, Minneapolis, MN. pp. 32-36.
  Sundaraj T, Kuruchew V, and Jayaraj J. 1998. Fungitoxicity of plant products and animal faeces on mycelial growth and sclerotial production of Rhizoctonia solani. Plant Dis Res 13:70-72.
• Sharma I and Nanda GS. 2000. Effect of plant extract on teliospores germination of Neovossia indica. Ind Phytopath 53(3):323-324.

DIRECTORATE OF WHEAT RESEARCH
Post Box 158, Agrasain Marg, Karnal 132001, India.

Facilitating the approach of participatory plant breeding to increase wheat yields in east and far-east regions of India. [p. 61-64]

Jag Shoran, Gyanendra Singh, B.S. Tyagi, Ravish Chatrath, Divakar Rai, Sarvan Kumar, and Surendra Singh.

Introduction. Wheat occupies a prime position in terms of production among the food crops in the world. In India, wheat is the second most important cereal crop and plays an important role in the food and nutritional security system of our country. Wheat alone contributes approximately a 25 % share of the total food grain production of the country. Wheat is consumed primarily in the form of an unleavened, pan-backed bread called chapati. Four wheat species, T. aestivum subsp. aestivum, T. turgidum subsp. durum, T. turgidum subsp. dicoccum, and T. aestivum subsp. sphaerococcum, are cultivated and consumed in one or the other form. In India, common bread wheat is the main crop, which occupies approximately 88 % of the area followed by T. turgidum subsp. durum (10 %) and T. turgidum subsp. dicoccum (2 %). Statistics from 2003-04 indicate the area under wheat cultivation to be approximately 27 million ha with a total production over 70 million tons. In terms of production and acreage, India is second to China among the wheat-growing countries in the world.

Wheat research and the yield gap in Indo-Gangetic Plains. Wheat improvement work in India began in 1905 when systematic research efforts were initiated with a series of selections from local types followed by pure-line selection that resulted in to the development of several better yielding, disease resistant, and quality wheats such as NP 4 and NP 6. With the inception of the All India Coordinated Wheat Improvement Project (AICWIP) in 1965, more than 200 wheat cultivars have been released in India for cultivation under various agroclimatic and production conditions. The results obtained from frontline demonstrations have shown an apparent yield gap in different agroclimatic conditions to the tune of 1.5 t/ha. The North Eastern Plains Zone, comprised of eastern UP, Bihar, Jharkhand, West Bengal, and Assam provinces, has about 9 million ha area under wheat with approximately 2.9 t/ha productivity compared to 4.1 t/ha in the North Western Plains Zone. Many improved, high-yielding genotypes have been released for different production conditions of North Eastern Plains Zone but very few could percolate to the farmers' fields. A myriad of reasons have been highlighted time and again.

The need for Participatory Varietal Selection (PVS) and Participatory Plant Breeding (PPB). In order to ensure the percolation of cultivars to the real beneficiary, a number of approaches have been suggested. Recently, the PVS and PPB approaches have been utilized to make wide adoption of improved cultivars and bridge the gap between potential and realized yields. PPB involves the plant breeder and farmers/clients in plant-breeding research and has been suggested as an effective alternative to formal plant breeding as a strategy for achieving productivity gains under low-input conditions. PPB is informal, with the involvement of farmer in helping plant breeders to develop plant ideotypes and also to provide feedback of farmers' preferences, helping decision making about the development and release of cultivars and seed production. In PPB, farmers take part in the dialogue regarding desirable plant characteristics, their presence or absence in specific genotypes, and also the traits farmers would like to see introduced. The farmers in this process are involved from the very beginning of plant-breeding programs that involves eliciting farmer's criteria for ranking alternative materials or contrasting plant characteristics in order of preference and then searching for parents, which offer some of the desired traits. PPB needs to be used only when the possibility of utilizing conventional and PVS approaches have been exhausted and when the search process fails to identify any suitable cultivars for commercial production in various microclimatic conditions. PPB also can exploit the results of PVS by using identified cultivars as parents of crosses. Compared to PVS/PPB, the increase in biodiversity will be at the intra- and intervarietal level and the effects of PPB will be more uneven than those of PVS, because the potential increase in the genetic diversity within a village is extremely large, whereas the increase in diversity with PVS is limited due to the limited cultivar diversity, i.e., only in the range of few cultivar choices.

Objectives, traits, and target area of PPB in India. The primary objective of PPB is tailoring genotypes to the specific micro agroclimatic and production conditions in such a manner that we develop a genotype for target environment rather than changing the environment. Defining target-area conditions, farmers' needs, and the environment are important. The collective efforts of researchers and farmers must address the farmer preferred traits and the target-area requirements through the alternate breeding approach of PPB. These two aspects in fact are prerequisites for initiating any PPB concept so as to define both traits and environment for their area of jurisdiction or stations that are further refined after a feedback through farmer appraisal and baseline surveys. In general, the target population of traits in the this particular area for which the program is designed represents a rainfed, limited irrigation condition, where drought and heat stress, short duration, and susceptibility to diseases (leaf blight and brown rust) were the major constraints. However, problem of high temperature coupled with high humidity causing a high incidence of leaf blight and spike sterility also are prevalent in high rainfall areas such as Assam and parts of West Bengal. The generalized traits that are well defined include early maturity, drought and heat tolerance, disease resistance, and bold amber grains with good chapati-making quality. A slight problem with preharvest sprouting, shattering, and late spike sterility is found in some parts of the target areas.

Keeping all above factors in view, we initiated a multipronged strategy to deal with the problems affecting the wheat yields directly or indirectly and thus to give an impetus to the wheat improvement program in the target areas of Ranchi and Assam where a sound breeding program is lacking due to various reasons. The Directorate of Wheat Research, Karnal, in close collaboration with the CIMMYT South Asia Regional Office, Kathmandu, Nepal, formulated and initiated a Department for International Development (DFID) funded project on entitled 'Participatory Research to Increase the Productivity and Sustainability of Wheat Cropping System in the State of Haryana, India'. The work on material development for facilitating the participatory plant breeding approach to be carried out at Shillongani and Ranchi Centre, was assigned to DWR Karnal. Accordingly, a base line survey was conducted in the target area to know the farmers' preference for the traits in a new cultivar. As anticipated and visualized through our experience of work in target area, total grain yield was ranked as the number one preference by the farmers, followed by a combination of yield, bold grain, and good chapati-making quality. A summary of the farmers preferred traits and their combinations are in Table 1.

Targeted crosses under PPB program. A set of seven diverse and promising cultivars with desired traits were selected for attempting the targeted crosses as per the need of both stations. The parental lines included at least one of the promising cultivars from North Eastern Plains Zone (DBW 14, HUW 468, PBW 443, and HUW 533), North Western Plains Zone (PBW 502), and Central Zone (DL 788-2 and GW 273). The maturity duration of the selected lines ranged from 106-142 days; yield potential was above 50 q/ha for irrigated conditions. During cultivar selection, disease flora and fauna were assessed and all cultivars were resistant to the brown rust and tolerant to leaf blight (Table 2). Hybridizations were primarily focused at improving the agronomic base to enhance yield levels and the acceptance of cultivars in the areas.

Planning, executing, and sharing of the crosses was implemented by DWR, Karnal. The F1 seed, along with the parents were planted in an off-season nursery in Dalang Maidan (10,000 feet above mean sea level) in the Lahaul and Spiti districts of Himachal Pradesh for generation advancement and evaluation. With the need for F2 material seed in mind for some important crosses, up to 800 F1 seeds were sent for multiplication in the off-season nursery. Details on the material and information generated during last two crop seasons are presented in Table 3.

The F2 material was supplied to both the centers namely Ranchi and Shillongani for their use as PPB base material. In addition, a set of 95 advanced bulks from selected crosses in the F6 and F7 generations also was evaluated at DWR during 2002-03 crop season. Out of this evaluation, 25 promising bulks were selected based on yield, maturity duration, plant height, and disease reactions were multiplied and shared with the Ranchi and Shillongani centers during the 2003-04 crop season to support the activities of PPB. Results on the performance of material under the target area suited to local needs are being utilized for fine-tuning the program for further enhancing the yield levels in east and far-east regions of India.

Acknowledgment. The authors are grateful to Dr. G.O. Ferrara, CIMMYT, SARO, Kathmandu, Nepal, and the DIFD Agency for technical and financial support provided for the present study. We also thankfully acknowledge the support from the coöperators in India.

References.

• Farringtion J and Martin A. 1988. Farmer participation in agricultural research: a review of concepts and practices. Agric Admin Unit Occasional Paper No. 9. ODI, London, UK.
• Friis-Hansen E. 1995. The role of local plant genetic resource management in participatory breeding. In: Proc Workshop on Participatory Plant Breeding, 26-29 July, 1995. Pp. 66-76.
• Harris D, Raghuwanshi BS, Gangwar JS, Singh SC, Joshi KD, Rashid A, and Hollington PA. 2001. Participatory evaluation by farmers of on-farm seed priming in wheat in India, Nepal and Pakistan. Exp Agric 37:403-415.
• Jacqueline A, Ashby T, Gracia MP, Guerrero C, Arturo Q, Roa JI, and Beltran JA. 1995. Innovation in the Organization of Participatory Plant Breeding. In: Proc Workshop on Participatory Plant Breeding, 26-29 July, 1995. Pp. 77-97.
• Joshi A and Witcombe JR. 1996. Farmer participatory crop improvement. II. Participatory varietal selection, a case study in India. Exp Agric 32:461-477.
• Jag S, Chatrath R, Singh G, Singh R, Tripathi SC, Sharma AK, Tyagi BS, and Singh SK. 2003. Participatory Research to Increase the Productivity and Sustainability of Wheat Cropping System in the State of Haryana, India. Ann Prog Rep Rev Plan Workshop, 10-14 June, 2003. CIMMYT, SARO, Nepal.
• Jag S, Chatrath R, Singh G, Singh R, Tripathi SC, Sharma AK, Tyagi BS, and Singh SK. 2004. Participatory Research to Increase the Productivity and Sustainability of Wheat Cropping System in the State of Haryana, India. Ann Prog Rep Rev Plan Workshop, 14-19 June, 2004. CIMMYT, SARO, Nepal.
• Singh G, Jag S, Tyagi BS, Chatrath R, Tripathi SC, and Nagarajan S. 2004. Participatory varietal selection in wheat, an approach to increase the adoption of new technologies. Dept Wheat Res, Karnal Res Bull No. 17 pp 28.
  Sthapit BR, Joshi KD, and Witcombe JR. 1996. Farmer participatory crop improvement. III. Participatory plant breeding, a case study for rice in Nepal. Exp Agric 32:479-496.
• Witcombe JR and Joshi A. 1995. The impact of farmer participatory research on biodiversity of crops. In: Proc Conf Using diversity and maintaining genetic resources on farm, New Delhi, June. IDRC.
• Witcombe JR, Joshi A, Joshi KD, and Sthapit BR. 1996. Farmer participatory crop improvement. I. Varietal selection and breeding method and their impact on biodiversity. Exp Agric 32:445-460.

Induction of variability through chemical mutagenesis in wheat. [p. 64-66]

S.K. Singh.

Wheat productivity in India has reached the saturation level because of the intensive use of available gene pool material in breeding programs. Mutation techniques are a novel approach for enhancing the level of genetic variability of a species within a short time. Selection can isolate superior genotypes (mutants) for various traits. Investigations on the effects of chemical mutagens for inducing variability have received much attention because of their importance in plant breeding. Among chemical mutagens, ethyl methane sulphonate (EMS) and sodium azide (SA) were used for inducing mutations in cereals (Awan et al. 1980; Georgiev 1982). Our experiment aimed to isolate and characterize mutants for yield traits using EMS and SA.

Approximately 1,000 healthy seeds of four high-yielding wheat genotypes, HP1633, HP1731, K9006, and K9107, were presoaked in distilled water for 1 h and treated in separate sets containing 0.01, 0.02, 0.03, and 0.04 M EMS (pH 7.0) and 0.5, 1.0, 1.5, and 2.0 mM SA (pH 3.0) prepared in fresh phosphate buffer. The seeds were completely submerged in the solutions (500 ml) for 4 h and then washed thoroughly in running water for 2 h before sowing to remove the residual chemicals. One thousand untreated dry seeds of all the four genotypes, soaked in distilled water for 4 h, served as the control.

A total of 36 treatment combinations including four controls were sown immediately after treatment with EMS and SA in Rabi 1995-96 at the Agriculture Research Farm, Banaras Hindu University, Varanasi. The plot size was 20 rows of 5 m with inter and intrarow spacing of 25 cm and 10 cm, respectively. All plants of each treatment representing M1 generation were harvested singly for producing an M2 generation. Seeds from individual M1 plants were space planted in a single 5-m row. Untreated seeds (control) also were sown after each 10th row for comparison. Individual plants were observed for various yield traits, and nine plants showing wide differences were selected and harvested separately to give the M3 generation. Mutants were confirmed as true breeding, because all mutant seeds yielded morphologically similar plants in the M3 that were quite distinct from the control. These mutants were harvested and planted in randomized block design with three replications in a double-row plot of 5 m in length in the crop season 1998-99. Ten plants from each mutant progeny row were used for characterization of the mutants for plant height (cm), number of tillers/plant, openness of flower glumes (degree), ear length (cm), number of grains/spike, 100-seed weight (g), yield/plant (g), grain shining, ear position, and lodging.

The data were subjected to AONVA (Panse and Sukhatme 1967). For yield traits, ANOVA using the mutant and control populations indicated all the treatments differed significantly for plant height, number of tillers/plant, openness of floret, spike length, number of grains/spike, 100-seed weight, and yield/plant.

Characterization of mutants. Nine mutants, superior for yield traits, were subjected to various doses of EMS and SA (Table 4). Both EMS and SA were effective in inducing variability in wheat genotypes at low and high concentrations of the chemicals, depending on the sensitivity of the genotype to the chemical and the concentration. The mutants were isolated and characterized for ten traits. WM1 (wheat mutant 1) was derived from the parent HP1633. WM2 and WM3 were mutants of HP1731. Chemical mutagenesis of K9006 produced mutants WM4, WM5, and WH6. WM7, WM8, and WM9 were isolated from the progenies of K9107.

Wheat mutant 1 (WM1). This mutant is high-yielding and bold seeded, with relatively higher number of grains/spike. WM1 had a significantly longer spike and wider angle of openness of flower than the control.

Wheat mutant 2 (WM2). A high-yielding mutant of HP1731 that had shiny, bold grains, a longer spike, and more grains/spike. WM2 also had a significantly wider angle of openness of flower than the control.

Wheat mutant 3 (WM3). This mutant was dwarf and a wider openness of glumes of the florets, but WM3 is poor for other traits.

Wheat mutant 4 (WM4). WM4 was a dwarf, high-yielding mutant having bold and shiny grains. This mutant also had significantly longer spikes and more profuse tillering than the parent K9006.

Wheat mutant 5 (WM5). This dwarf mutant was high-yielding and long-spiked with shiny grains and profuse tillering. WM5 had more seeds/spike and wider openness of glumes of the florets. This mutant also had a high stem strength, indicating its superiority for lodging resistance.

Wheat mutant 6 (WM6). This dwarf and high-yielding mutant possessed long, erect spikes with wider openness of glumes of the florets.

Wheat mutant 7 (WM7). WM7 was a high-yielding mutant having shiny, bold grains. WM7 also was for number of seeds/spike and openness of florets over the parent K9107.

Wheat mutant 8 (WM8). This dwarf mutant had significantly more seeds/spike and openness of florets compared to the K9107 parent.

Wheat mutant 9(WM9). WM9, also a dwarf mutant, had profuse tillering and more grains/spike, and a wider openness of the florets.

In general, most of the mutants were high yielding compared to parent cultivars and can be effectively utilized in hybrid-development program because of their wider openness of glumes of the florets, which is expected to promote out crossing.

References.

• Panse VG and Sukhatme PV. 1967. Statistical methods for agricultural workers, ICAR Publication, New Delhi.
• Awan MA, Konzak CF, Nilan JN, and Nilan RA. 1980. Mutagenic effects of sodium azide in rice. Crop Sci 20:663-668.
• Georgiev SA. 1982. Male sterile mutants induced in T. aestivum after EMS treatment. Comptes Rendus de 1' Academie Bulgare des Sciences 35:241-243.

Conservation of wheat germ plasm under natural conditions in the Lahul-Spiti Valley in the Himalayan Hills of India. [p. 66-67]

S.K. Singh and S. Kundu.

The germination of 30 randomly selected wheat accessions from the wheat germ plasm maintained in different types of packing under natural conditions in Lahul Spiti, Himanchal Pradesh, was tested for 4 consecutive years. We observed that the seeds stored in aluminum envelopes had more than an 88 % germination rate and, thus, seeds can be maintained up to years in this type of packaging in a cost-effective manner.

Genetic resources are the prerequisite for a successful breeding program. India has achieved a tremendous jump in wheat production and ranks second worldwide, a result of using indigenous and exotic gene pools extensively in breeding programs. At present, conserving all the available germ plasm is a necessity. Conservation of these accessions requires a highly specific setup for maintaining the seed viability for long periods. Sixty-eight released cultivars were sent for conservation under natural condition (cold, dry environment) at the Wheat Summer Nursery (WSN) Facility at Dalang Maidan in the Lahul-Spiti Valley of the Himanchal Pradesh state for the first time in 1998. This station is located in the Himalayan Hills Range at more than 10,000 ft above mean sea level. The germination percent of the cultivars ranged from 98 % to 100 %. An experiment tested the effect of the prevailing environment on germination of wheat seeds conserved in different packaging over the years.

Of these 68 accessions, 30 (24 T. aestivum subsp. aestivum, four T. turgidum subsp. durum, one T. turgidum subsp. dicoccum, and one triticale) were selected randomly for the study. The seeds were stored in three types of packaging, cloth bags, waterproof envelopes, and aluminum envelopes. Beginning in 1999, seeds of these 30 accessions were removed from all packaging and germination tested over four consecutive years (1999-2002). ANOVA (Panse and Sukhatme 1967) was done and critical differences were used to compare germination in different packaging and duration of conservation.

ANOVA indicated the highly significant differences for the various packaging methods, year, and the 'packaging x year' interaction. Results on germination percent of the genotypes each year indicated a significant reduction over the years compared to first year but remained good during the first 3 years of their storage under natural conditions in the Lahul Valley. A drastic reduction in germination wa