Harcharan Singh Dhaliwal

Mapping of Quantitative Trait Loci for Grain Iron and Zinc Concentration in Diploid A Genome Wheat

Micronutrients, especially iron (Fe) and zinc (Zn), are deficient in the diets of people in underdeveloped countries. Biofortification of food crops is the best approach for alleviating the micronutrient deficiencies. Identification of germplasm with high grain Fe and Zn and understanding the genetic basis of their accumulation are the prerequisites for manipulation of these micronutrients. Some wild relatives of wheat were found to have higher grain Fe and Zn concentrations compared with the cultivated bread wheat germplasm. One accession of Triticum boeoticum (pau5088) that had relatively higher grain Fe and Zn was crossed with Triticum monococcum (pau14087), and a recombinant inbred line (RIL) population generated from this cross was grown at 2 locations over 2 years. The grains of the RIL population were evaluated for Fe and Zn concentration using atomic absorption spectrophotometer. The grain Fe and Zn concentrations in the RIL population ranged from 17.8 to 69.7 and 19.9 to 64.2 mg/kg, respectively. A linkage map available for the population was used for mapping quantitative trait loci (QTL) for grain Fe and Zn accumulation. The QTL analysis led to identification of 2 QTL for grain Fe on chromosomes 2A and 7A and 1 QTL for grain Zn on chromosome 7A. The grain Fe QTL were mapped in marker interval Xwmc382-Xbarc124 and Xgwm473-Xbarc29, respectively, each explaining 12.6% and 11.7% of the total phenotypic variation and were designated as QFe.pau-2A and QFe.pau-7A. The QTL for grain Zn, which mapped in marker interval Xcfd31-Xcfa2049, was designated as QZn.pau-7A and explained 18.8% of the total phenotypic variation.

Identification and mapping of a tiller inhibition gene (tin3) in wheat

Tillering is one of the most important agronomic traits in cereal crops because tiller number per plant determines the number of spikes or panicles per plant, a key component of grain yield and/or biomass. In order to characterize the underlying genetic variation for tillering, we have isolated mutants that are compromised in tillering ability using ethyl methanesulphonate (EMS)-based mutagenesis in diploid wheat (Triticum monococcum subsp. monococcum). The tillering mutant, tiller inhibition (tin3) produces only one main culm compared to the wild type with many tillers. The monoculm phenotype of tin3 is due to a single recessive mutation. Genetic and molecular mapping in an F2 population of diploid wheat located the tin3 gene on the long arm of chromosome 3Am. One codominant RFLP marker Xpsr1205 cosegregated with tin3 in the F2 population. Physical mapping of PSR1205 in a set of Chinese Spring deletion lines of group-3 chromosomes placed the tin3 gene in the distal 10% of the long arm of chromosome 3A, which is a recombination-rich region in wheat. The implications of the mapping of tin3 on chromosome arm 3AmL are discussed with respect to putative orthologs of tin3 in the 3L colinear regions across various cereal genomes and other tillering traits in grasses.

Pyramiding of two bacterial blight resistance and a semidwarfing gene in Type 3 Basmati using marker-assisted selection

A traditional Type 3 Basmati rice cultivar grown in India is tall and lodges even under low nitrogen fertilizer dose. In addition to lodging, it is highly susceptible to several diseases and pests including bacterial blight (BB). BB resistance genes (Xa21 and xa13) and a semidwarfing gene (sd-1) were pyramided in Type 3 Basmati from a rice cultivar PR106-P2 using marker-assisted selection (MAS). Foreground selection for BB resistance genes, Xa21 and xa13 and reduced plant height gene, sd-1 was carried on the basis of linked molecular markers pTA248, RG136 and ‘h’, respectively. The BC2F3 progenies with both the BB resistance genes were highly resistant with lower lesion length than either of the genes individually. Background profiling of the selected 16 BC2F3 progenies was done using 95 anchored SSR and 12 ISSR markers. Among the selected 16 BC2F3 progenies, 38-5-2 and 38-5-36 closely clustered along with the recipient parent Type 3 Basmati showing above 85% genetic similarity with the same. Further selection was continued till F5 generation for higher recovery for Type 3 Basmati characteristics. The desirable alleles of intermediate amylose content (wx) and aroma (fgr) loci of Type 3 Basmati were also tracked using the linked SSR markers. The BC2F5 pyramid lines T3-4, T3-5, T3-6 and T3-7 homozygous for the three target genes Xa21, xa13 and sd-1 from the donor parent with wx and fgr alleles of Type 3 Basmati had excellent cooking quality and strong aroma.

Development of Triticum turgidum subsp. durum – Aegilops longissima amphiploids with high iron and zinc content through unreduced gamete formation in F1 hybrids

Four different interspecific hybrids involving three different accessions of Aegilops longissima Schweinf. & Muschl. with high grain iron and zinc content and three Triticum turgidum L. subsp. durum (Desf.) Husn. cultivars with low micronutrient content were made for durum wheat biofortification and investigated for chromosome pairing, fertility, putative amphiploidy, and micronutrient content. The chromosome pairing in the 21-chromosome F1 hybrids (ABSl) consisted of 0-6 rod bivalents and occasionally 1 trivalent. All the F1 hybrids, however, unexpectedly showed partial but variable fertility. The detailed meiotic investigation indicated the simultaneous occurrence of two types of aberrant meiotic divisions, namely first-division restitution and single-division meiosis, leading to regular dyads and unreduced gamete formation and fertility. The F2 seeds, being putative amphiploids (AABBSlSl), had nearly double the chromosome number (40-42) and regular meiosis and fertility. The F1 hybrids were intermediate between the two parents for different morphological traits. The putative amphiploids with bold seed size had higher grain ash content and ash iron and zinc content than durum wheat cultivars, suggesting that Ae. longissima possesses a better genetic system(s) for uptake and seed sequestration of iron and zinc, which could be transferred to elite durum and bread wheat cultivars and exploited.

Transfer of leaf rust and stripe rust resistance from Aegilops umbellulata Zhuk. to bread wheat (Triticum aestivum L.)

Aegilops umbellulata acc. 3732, an excellent source of resistance to major wheat diseases, was used for transferring leaf rust and stripe rust resistance to cultivated wheat. An amphiploid between Ae. umbellulata acc. 3732 and Triticum durum cv. WH890 was crossed with cv. Chinese Spring PhI to induce homoeologous pairing between Ae. umbellulata and wheat chromosomes. The F1 was crossed to the susceptible Triticum aestivum cv. ‘WL711’ and leaf rust and stripe rust resistant plants were selected among the backcross progenies. Homozygous lines were selected and screened against six Puccinia triticina and four Puccinia striiformis f. sp. tritici pathotypes at the seedling stage and a mixture of prevalent pathotypes of both rust pathogens at the adult plant stage. Genomic in situ hybridization in some of the selected introgression lines detected two lines with complete Ae. umbellulata chromosomes. Depending on the rust reactions and allelism tests, the introgression lines could be classified into two groups, comprising of lines with seedling leaf rust resistance gene Lr9 and with new seedling leaf rust and stripe rust resistance genes. Inheritance studies detected an additional adult plant leaf rust resistance gene in one of the introgression lines. A minimum of three putatively new genes—two for leaf rust resistance (LrU1 and LrU2) and one for stripe rust resistance (YrU1) have been introgressed into wheat from Ae. umbellulata. Two lines with no apparent linkage drag have been identified. These lines could serve as sources of resistance to leaf rust and stripe rust in breeding programs.

Bioavailability of Iron from Wheat Aegilops Derivatives Selected for High Grain Iron and Protein Contents

A coupled in vitro digestion/Caco-2 model was employed to assess iron bioavailability from wheat Aegilops derivatives selected for high iron and protein contents. The iron content in wheat genotypes used in this study correlated to a great extent with both protein (r = 0.80) and phytate (r = 0.68) contents. The iron bioavailability was based on Caco-2 cell ferritin formation from cooked digests of these derivatives (relative to WL711 control) and correlated positively with dialyzable iron (r = 0.63) and total iron content (r = 0.38) but not with the phytate content. The apparently decreased phytate/iron molar ratios, however, correlated negatively (r = -0.42) with the iron bioavailability, justifying the utilization of these parameters in biofortification programs. Iron bioavailability in the derivatives increased up to 1.5-fold, corresponding to a 1.5-2.2-fold increase observed in iron content over control. These data suggest that biofortification for iron proportionately leading to higher iron bioavailability will be the most feasible and cost-effective approach to combat micronutrient deficiency.

Molecular mapping of cereal cyst nematode resistance in Triticum monococcum L. and its transfer to the genetic background of cultivated wheat

Triticum monococcum, the diploid A genome species, harbours enormous variability for resistance to biotic stresses. A spring type T. monococcum acc. 14087 was found to be resistant to Heterodera avenae (cereal cyst nematode, CCN). A recombinant inbred line population (RIL) developed by crossing this accession with a CCN susceptible T. boeoticum acc. 5088 was used for studying the inheritance and map location of the CCN resistance. Based on composite interval mapping two QTL, one each on chromosome 1AS and 2AS, were detected. The QTL on 1A, designated as Qcre.pau-1A, appeared to be a major gene with 26% contribution to the overall phenotypic variance whereas the QTL on 2A designated as Qcre.pau-2A contributed 13% to total phenotypic variation. Qcre.pau-1A is novel, being the only CCN resistance gene mapped in any ‘A’ genome species and none of the other known genes have been mapped on chromosome 1A. The QTL Qcre.pau-2A might be allelic to Cre5, a CCN resistance gene transferred from Ae. ventricosa and mapped on 2AS. The Qcre.pau-1A was transferred to cultivated wheat using T. durum cv. PBW114 as the bridging species. Selected CCN resistant F8 lines showed introgression for the molecular markers identified to be linked with CCN resistance locus Qcre.pau-1A. Thus, this gene alone could impart complete resistance against CCN. These introgression lines can be used for marker-assisted transfer of Qcre.pau-1A to elite wheat cultivars.

Development and characterization of Triticum aestivum – Aegilops kotschyi amphiploids with high grain iron and zinc contents

Synthetic amphiploids between Triticum aestivum (AABBDD) landrace Chinese Spring ( Ph I ) and cultivar WL711 with different accessions of Aegilops kotschyi (UUS l S l ) were developed through colchicine treatment of sterile hybrids. The F 1 hybrids and amphiploid plants were intermediate between the parents for plant morphology and spike characteristics. Meiotic metaphase chromosome analysis of the F 1 hybrids (ABDUS l ) showed the expected chromosome number (35) and very little but variable homoeologous chromosome pairing. The amphiploids (AABBDDUUS l S l ), however, had variable frequency of univalents at meiotic metaphase-I. The SDS–PAGE of high molecular weight glutenin subunits of amphiploids along with the parents showed the presence and expression of all the parental genomes in the amphiploids. The amphiploids with seeds as large as that of wheat cultivars had higher grain, flag leaf and grain ash iron and zinc concentrations than the wheat parents and comparable with those of their Ae. kotschyi parents suggest that Ae. kotschyi possesses a distinctive genetic system for the micronutrient uptake, translocation and sequestration than the wheat cultivars. This could, however, be demonstrated unequivocally only with comprehensive data on biomass, grain yield and harvest index of the Aegilops donors and the synthetic amphiploids, which is not feasible due to their shattering and hard threshing. The use of amphiploids for the transfer of high iron and zinc concentrations and development of alien addition and substitution lines in wheat is in progress.

Food waste: a potential bioresource for extraction of nutraceuticals and bioactive compounds

Food waste, a by-product of various industrial, agricultural, household and other food sector activities, is rising continuously due to increase in such activities. Various studies have indicated that different kind of food wastes obtained from fruits, vegetables, cereal and other food processing industries can be used as potential source of bioactive compounds and nutraceuticals which has significant application in treating various ailments. Different secondary metabolites, minerals and vitamins have been extracted from food waste, using various extraction approaches. In the next few years these approaches could provide an innovative approach to increase the production of specific compounds for use as nutraceuticals or as ingredients in the design of functional foods. In this review a comprehensive study of various techniques for extraction of bioactive components citing successful research work have been discussed. Further, their efficient utilization in development of nutraceutical products, health benefits, bioprocess development and value addition of food waste resources has also been discussed.

A candidate gene OsAPC6 of anaphase-promoting complex of rice identified through T-DNA insertion

A dwarf mutant (Oryza sativa anaphase-promoting complex 6 (OsAPC6)) of rice cultivar Basmati 370 with 50% reduced plant height as compared to the wild type was isolated by Agrobacterium tumefaciens-mediated transformation using HmRDs cassette. This mutant was found to be insensitive to exogenous gibberellic acid (GA3) application. Homozygous mutant plants showed incomplete penetrance and variable expressivity for plant height and pleiotropic effects including gibberellic acid insensitivity, reduced seed size, panicle length, and female fertility. Single copy insertion of T-DNA and its association with OsAPC6 was confirmed by Southern hybridization, germination on hygromycin, and 3:1 segregation of HPT gene in F2 from OsAPC6 × Basmati 370 cross. The T-DNA flanking region sequenced through thermal asymmetric interlaced polymerase chain reaction showed a single hit on chromosome 3 of japonica rice cultivar Nipponbare in the second exonic region of a gene which encodes for sixth subunit of anaphase-promoting complex/cyclosome. The candidate gene of 8.6-kb length encodes a 728-amino acid protein containing a conserved tetratricopeptide repeat (TPR) domain and has only a paralog, isopenicillin N-synthase family protein on the same chromosome without the TPR domain. There was no expression of the gene in the mutant while in Basmati 370, it was equal in both roots and shoots. The knockout mutant OsAPC6 interferes with the gibberellic acid signaling pathway leading to reduced height and cell size probably through ubiquitin-mediated proteolysis. Further functional validation of the gene through RNAi is in progress.