Mining of Indian wheat germplasm collection for adult plant resistance to leaf rust.

Leaf rust (Puccinia triticina Eriks.) is a fungal disease of wheat (Triticum spp.), which causes considerable yield loss. Adult plant resistance (APR) is one of the most sustainable approaches to control leaf rust. In this study, field-testing was carried out across ten different locations, followed by molecular screening, to detect the presence of APR genes, Lr34+, Lr46+, Lr67+ and Lr68 in Indian wheat germplasm. In field screening, 190 wheat accessions were selected from 6,319 accessions based on leaf tip necrosis (LTN), disease severity and the average coefficient of infection. Molecular screening revealed that 73% of the accessions possessed known APR genes either as single or as a combination of two or three genes. The occurrence of increased LTN intensity, decreased leaf rust severity and greater expression of APR genes were more in relatively cooler locations. In 52 lines, although the presence of the APR genes was not detected, it still displayed high levels of resistance. Furthermore, 49 accessions possessing either two or three APR genes were evaluated for stability across locations for grain yield. It emerged that eight accessions had wider adaptability. Resistance based on APR genes, in the background of high yielding cultivars, is expected to provide a high level of race non-specific resistance, which is durable.

Identification and characterization of finger millet OPAQUE2 transcription factor gene under different nitrogen inputs for understanding their role during accumulation of prolamin seed storage protein.

In this study, we report the isolation and characterization of the mRNA encoding OPAQUE2 (O2) like TF of finger millet (FM) (Eleusine coracana) (EcO2). Full-length EcO2 mRNA was isolated using conserved primers designed by aligning O2 mRNAs of different cereals followed by 3' and 5' RACE (Rapid Amplification of cDNA Ends). The assembled full-length EcO2 mRNA was found to contain an ORF of 1248-nt coding the 416 amino acids O2 protein. Domain analysis revealed the presence of the BLZ and bZIP-C domains which is a characteristic feature of O2 proteins. Phylogenetic analysis of EcO2 protein with other bZIP proteins identified using finger millet transcriptome data and O2 proteins of other cereals showed that EcO2 shared high sequence similarity with barley BLZ1 protein. Transcripts of EcO2 were detected in root, stem, leaves, and seed development stages. Furthermore, to investigate nitrogen responsiveness and the role of EcO2 in regulating seed storage protein gene expression, the expression profiles of EcO2 along with an α-prolamin gene were studied during the seed development stages of two FM genotypes (GE-3885 and GE-1437) differing in grain protein content (13.8 and 6.2%, respectively) grown under increasing nitrogen inputs. Compared to GE-1437, the EcO2 was relatively highly expressed during the S2 stage of seed development which further increased as nitrogen input was increased. The Ecα-prolamin gene was strongly induced in the high protein genotype (GE-3885) at all nitrogen inputs. These results indicate the presence of nitrogen responsiveness regulatory elements which might play an important role in accumulating protein in FM genotypes through modulating EcO2 expression by sensing plant nitrogen status.

Genotyping-by-Sequencing Analysis for Determining Population Structure of Finger Millet Germplasm of Diverse Origins.

Finger millet [ (L.) Gaertn.] is grown mainly by subsistence farmers in arid and semiarid regions of the world. To broaden its genetic base and to boost its production, it is of paramount importance to characterize and genotype the diverse gene pool of this important food and nutritional security crop. However, as a result of nonavailability of the genome sequence of finger millet, the progress could not be made in realizing the molecular basis of unique qualities of the crop. In the present investigation, attempts have been made to characterize the genetically diverse collection of 113 finger millet accessions through whole-genome genotyping-by-sequencing (GBS), which resulted in a genome-wide set of 23,000 single-nucleotide polymorphisms (SNPs) segregating across the entire collection and several thousand SNPs segregating within every accession. A model-based population structure analysis reveals the presence of three subpopulations among the finger millet accessions, which are in parallel with the results of phylogenetic analysis. The observed population structure is consistent with the hypothesis that finger millet was domesticated first in Africa, and from there it was introduced to India some 3000 yr ago. A total of 1128 gene ontology (GO) terms were assigned to SNP-carrying genes for three main categories: biological process, cellular component, and molecular function. Facilitated access to high-throughput genotyping and sequencing technologies are likely to improve the breeding process in developing countries, and as such, this data will be very useful to breeders who are working for the genetic improvement of finger millet.

Alteration of Genetic Make-up in Karnal Bunt Pathogen (Tilletia indica) of Wheat in Presence of Host Determinants.

Alteration of genetic make-up of the isolates and monosporidial strains of Tilletia indica causing Karnal bunt (KB) disease in wheat was analyzed using DNA markers and SDS-PAGE. The generation of new variation with different growth characteristics is not a generalized feature and is not only dependant on the original genetic make up of the base isolate/monosporidial strains but also on interaction with host. Host determinant(s) plays a significant role in the generation of variability and the effect is much pronounced in monosporidial strains with narrow genetic base as compared to broad genetic base. The most plausible explanation of genetic variation in presence of host determinant(s) are the recombination of genetic material from two different mycelial/sporidia through sexual mating as well as through para-sexual means. The morphological and development dependent variability further suggests that the variation in T. indica strains predominantly derived through the genetic rearrangements.

Association mapping of agro-morphological characters among the global collection of finger millet genotypes using genomic SSR markers.

Identification of alleles responsible for various agro-morphological characters is a major concern to further improve the finger millet germplasm. Forty-six genomic SSRs were used for genetic analysis and population structure analysis of a global collection of 190 finger millet genotypes and fifteen agro-morphological characters were evaluated. The overall results showed that Asian genotypes were smaller in height, smaller flag leaf length, less basal tiller number, early flowering and early maturity nature, small ear head length, and smaller in length of longest finger. The 46 SSRs yielded 90 scorable alleles and the polymorphism information content values varied from 0.292 to 0.703 at an average of 0.442. The gene diversity was in the range of 0.355 to 0.750 with an average value of 0.528. The 46 genomic SSR loci grouped the 190 finger millet genotypes into two major clusters based on their geographical origin by the both phylogenetic clustering and population structure analysis by STRUCTURE software. Association mapping of QTLs for 15 agro-morphological characters with 46 genomic SSRs resulted in identification of five markers were linked to QTLs of four traits at a significant threshold (P) level of ≤ 0.01 and ≤ 0.001. The QTL for basal tiller number was strongly associated with the locus UGEP81 at a P value of 0.001 by explaining the phenotypic variance (R (2)) of 10.8%. The QTL for days to 50% flowering was linked by two SSR loci UGEP77 and UGEP90, explained 10 and 8.7% of R (2) respectively at a P value of 0.01. The SSR marker, FM9 found to have strong association to two agro-morphological traits, flag leaf width (P-0.001, R(2)-14.1 %) and plant height (P-0.001, R(2)-11.2%). The markers linked to the QTLs for above agro-morphological characters found in the present study can be further used for cloning of the full length gene, fine mapping and their further use in the marker assisted breeding programmes for introgression of alleles into locally well adapted germplasm.

Cloning and characterization of cold, salt and drought inducible C-repeat binding factor gene from a highly cold adapted ecotype of Lepidium latifolium L.

The dehydration-responsive element-binding (DREB) protein/C-Repeat Binding Factors (CBFs) belongs to APETALA2 (AP2) family transcription factors that binds to DRE/CRT cis-element in cold-responsive (COR) genes and induce COR genes. CBFs have been isolated and characterized from evolutionarily diverse plant species. CBF pathway is conserved by CBF regulon and the size or the number and kind of target genes vary among freezing sensitive and tolerant plants. Hence, cloning of CBFs from highly freezing tolerant plants such as Lepidium latifolium L. will be useful in understanding the freezing tolerance of this species. In this study, LlCBF, a CBF1 family gene from L. Latifolium L., was cloned using RT-PCR and RACE-PCR. The full length mRNA of LlCBF is 948 bp with an open reading frame of 642 bp, encoding a protein of 213 amino acids with a molecular weight of 23.92 kDa and a theoretical isoelectric point of 4.80. Amino acid sequence analysis showed that LlCBF has an AP2 DNA binding domain, a potential CBF type nuclear localization signal (NLS) and C-terminal acidic domain. Semi-quantitative RT-PCR analysis of LlCBF revealed that this gene is up-regulated by high salt, dehydration and low temperature stresses. The investigation is therefore successful in cloning of a gene having strong homology with CBF transcription factors and responsive to low temperature, high salt and dehydration conditions.

DREB1/CBF transcription factors: their structure, function and role in abiotic stress tolerance in plants.

Drought, high salinity and low temperature are major abiotic stresses that influence survival, productivity and geographical distribution of many important crops across the globe. Plants respond to these environmental challenges via physiological, cellular and molecular processes, which results in adjusted metabolic and structural alterations. The dehydration-responsiveelement-binding (DREB) protein / C-repeat binding factors (CBFs) belong to APETALA2 (AP2) family transcription factors that bind to DRE/CRT cis-element and regulate the expression of stress-responsive genes. DREB1/CBF genes, therefore, play an important role in increasing stress tolerance in plants and their deployment using transgenic technology seems to be a potential alternative in management of abiotic stresses in crop plants. This review is mainly focussed on the structural characteristics as well as transcriptional regulation of gene expression in response to various abiotic stresses, with particular emphasis on the role of DREB1/CBF regulon in stress-responsive gene expression. The recent progress related to genetic engineering of DREB1/CBF transcription factors in various crops and model plants is also summarized.

Deep vein thrombosis in protein S deficiency.

Protein S is a vitamin K-dependent anticoagulant protein. It functions as a cofactor of activated protein C to inactivate activated factor V (FVa) and activated factor VIII (FVIIIa). Its deficiency is a rare condition and can lead to deep vein thrombosis, pulmonary embolism or stroke. It is often treated with long-term anti-coagulant therapy. Protein S deficiency may be hereditary or acquired; the latter is usually due to hepatic diseases or a vitamin K deficiency. Protein S deficiency manifests as an autosomal dominant trait; manifestations of thrombosis are observed in both heterozygous and homozygous genetic deficiencies of protein S. This case report is of DVT due to Protein S deficiency in a 53 year old male. Venous Doppler was used to diagnose DVT and free Protein S level measured by ELISA. IVC filter was placed on the third day of admission.

Significance of Cryptosporidium as an aetiology of acute infectious diarrhoea in elderly Indians.

OBJECTIVE: To explore the significance of Cryptosporidium isolation in elderly Indian with acute infectious diarrhoea and its clinical correlates. DESIGN: A hospital-based clinico-aetiological study of 120 patients aged 60 years or older, 25 adults younger than 60 years and 25 children up to age 14 with acute diarrhoea, and 57 apparently healthy elderly individuals. RESULTS: Cryptosporidium was isolated in 22 older patients with diarrhoea (18.3%) and was highly significant (P < 0.01) compared with healthy age-matched controls. Of these patients 66% had a history of close contact with animals. Most (68%) Cryptosporidium infections occurred during the rainy season. Among the elderly patients 17% suffered from vomiting and abdominal pain, 31% were febrile; none were severely dehydrated. Stools numbered three to nine per day with duration of 5-17 days. Stool leucocytes were <6/hpf and no RBCs were seen. Isolation of Cryptosporidium in older persons was associated with diabetes mellitus (22.7%), tuberculosis (9.0%), malignancy (4.5%) and coronary artery disease (4.5%). CONCLUSION: Cryptosporidium is an important cause of diarrhoea in elderly Indians, especially those with close contact with animals. The infection has a mild clinical course, is self-limiting and does not cause dysenteric stool. Its effect on nutrition requires further study.