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Not AvailableWheat is a staple food worldwide and provides 40% of the calories in the diet. Climate change and global warming pose a threat to wheat production, however, and demand a deeper understanding of how heat stress might impact wheat production and wheat biology. However, it is difficult to identify novel heat stress associated genes when the genomic information is not available. Wheat has a very large and complex genome that is about 37 times the size of the rice genome. The present study sequenced the whole transcriptome of the wheat cv. HD2329 at the flowering stage, under control (22°±3° C) and heat stress (42° C, 2 h) conditions using Illumina HiSeq and Roche GS-FLX 454 platforms. We assembled more than 26.3 and 25.6 million high-quality reads from the control and HS-treated tissues transcriptome sequences respectively.Not Availabl

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Not AvailableWheat is a staple food worldwide and provides 40% of the calories in the diet. Climate change and global warming pose a threat to wheat production, however, and demand a deeper understanding of how heat stress might impact wheat production and wheat biology. However, it is difficult to identify novel heat stress associated genes when the genomic information is not available. Wheat has a very large and complex genome that is about 37 times the size of the rice genome. The present study sequenced the whole transcriptome of the wheat cv. HD2329 at the flowering stage, under control (22 – 3C) and heat stress (42C, 2 h) conditions using Illumina HiSeq and Roche GS-FLX 454 platforms. We assembled more than 26.3 and 25.6 million high-quality reads from the control and HS-treated tissues transcriptome sequences respectively. About 76,556 (control) and 54,033 (HS-treated) contigs were assembled and annotated de novo using different assemblers and a total of 21,529 unigenes were obtained. Gene expression profile showed significant differential expression of 1525transcripts under heat stress, of which 27 transcripts showed very high (>10) fold upregulation. Cellular processes such as metabolic processes, protein phosphorylation, oxidations-reductions, among others were highly influenced by heat stress. In summary, these observations significantly enrich the transcript dataset of wheat available on public domain and show a de novo approach to discover the heat-responsive transcripts of wheat, which can accelerate the progress of wheat stress-genomics as well as the course of wheat breeding programs in the era of climate change.Not Availabl

Identification of putative RuBisCo activase (TaRca1) ˗ the catalytic chaperone regulating carbon assimilatory pathway in wheat (Triticum aestivum) under the heat stress

RuBisCo activase (Rca) is a catalytic chaperone involved in modulating the activity of RuBisCo (key enzyme of photosynthetic pathway). Here, we identified eight novel transcripts from wheat through data mining predicted to be Rca and cloned a transcript of 1.4 kb from cv. HD2985, named as TaRca1 (GenBank acc. no. KC776912). Single copy number of TaRca1 was observed in wheat genome. Expression analysis in diverse wheat genotypes (HD2985, Halna, PBW621 and HD2329) showed very high relative expression of TaRca1 in Halna under control and HS-treated, as compared to other cultivars at different stages of growth. TaRca1 protein was predicted to be chloroplast-localized with numerous potential phosphorylation sites. Nothern blot analysis showed maximum accumulation of TaRca1 transcript in thermotolerant cv. during mealy-ripe stage, as compared to thermosusceptible. Decrease in the photosynthetic parameters was observed in all the cultivars, except PBW621 in response to HS. We observed significant increase in the Rca activity in all the cultivars under HS at different stages of growth. HS causes decrease in the RuBisCo activity; maximum reduction was observed during pollination stage in thermosusceptible cvs. as validated through immunoblotting. We observed uniform carbon distribution in different tissues of thermotolerant cvs., as compared to thermosusceptible. Similarly, tolerance level of leaf was observed maximum in Halna having high Rca activity under HS. A positive correlation was observed between the transcript and activity of TaRca1 in HS-treated Halna. Similarly, TaRca1 enzyme showed positive correlation with the activity of RuBisCo. There is, however, need to manipulate the thermal stability of TaRca1 enzyme through protein engineering for sustaining the photosynthetic rate under HS – a novel approach towards development of ‘climate-smart’ crop

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Not AvailableRuBisCo activase (Rca) is a catalytic chaperone involved in modulating the activity of RuBisCo (key enzyme of photosynthetic pathway). Here, we identified eight novel transcripts from wheat through data mining predicted to be Rca and cloned a transcript of 1.4 kb from cv. HD2985, named as TaRca1 (GenBank acc. no. KC776912). Single copy number of TaRca1 was observed in wheat genome. Expression analysis in diverse wheat genotypes (HD2985, Halna, PBW621, and HD2329) showed very high relative expression of TaRca1 in Halna under control and HS-treated, as compared to other cultivars at different stages of growth. TaRca1 protein was predicted to be chloroplast-localized with numerous potential phosphorylation sites. Northern blot analysis showed maximum accumulation of TaRca1 transcript in thermotolerant cv. during mealy-ripe stage, as compared to thermosusceptible. Decrease in the photosynthetic parameters was observed in all the cultivars, except PBW621 in response to HS. We observed significant increase in the Rca activity in all the cultivars under HS at different stages of growth. HS causes decrease in the RuBisCo activity; maximum reduction was observed during pollination stage in thermosusceptible cvs. as validated through immunoblotting. We observed uniform carbon distribution in different tissues of thermotolerant cvs., as compared to thermosusceptible. Similarly, tolerance level of leaf was observed maximum in Halna having high Rca activity under HS. A positive correlation was observed between the transcript and activity of TaRca1 in HS-treated Halna. Similarly, TaRca1 enzyme showed positive correlation with the activity of RuBisCo. There is, however, need to manipulate the thermal stability of TaRca1 enzyme through protein engineering for sustaining the photosynthetic rate under HS—a novel approach toward development of “climate-smart” crop.Not Availabl

Outcomes of coronary artery bypass grafting and reduction annuloplasty for functional ischemic mitral regurgitation: A prospective multicenter study (Randomized Evaluation of a Surgical Treatment for Off-Pump Repair of the Mitral Valve)

ObjectiveFunctional ischemic mitral regurgitation is a complication of ventricular remodeling; standard therapy is reduction annuloplasty and coronary artery bypass grafting. Unfortunately, outcomes are retrospective and contradictory. We report a multicenter study that documents the outcomes of reduction annuloplasty for functional ischemic mitral regurgitation.MethodsTwenty-one centers randomized 75 patients to the coronary artery bypass grafting  +  reduction annuloplasty subgroup that was the control arm of the Randomized Evaluation of a Surgical Treatment for Off-pump Repair of the Mitral Valve trial. Entry criteria included patients requiring revascularization, patients with severe or symptomatic moderate functional ischemic mitral regurgitation, an ejection fraction 25% or greater, a left ventricular end-diastolic dimension 7.0 cm or less, and more than 30 days since acute myocardial infarction. All echocardiograms were independently scored by a core laboratory. Reduction annuloplasty was achieved by device annuloplasty. Two patients underwent immediate intraoperative conversion to a valve replacement because reduction annuloplasty was unable to correct mitral regurgitation; as-treated results are presented.ResultsThirty-day mortality was 4.1% (3/73). Patients received an average of 2.8 bypass grafts. Mean follow-up was 24.6 months. Mitral regurgitation was reduced from 2.6 ± 0.8 preoperatively to 0.3 ± 0.6 at 2 years. Freedom from death or valve reoperation was 78% ± 5% at 2 years. There was significant improvement in ejection fraction and New York Heart Association class with reduction of left ventricular end-diastolic dimension. Cox regression analyses suggested that increasing age (P = .001; hazard ratio, 1.16 per year; 95% confidence interval, 1.06–1.26) and renal disease (P = .018; hazard ratio, 3.48; 95% confidence interval, 1.25–9.72) were associated with decreased survival.ConclusionsCoronary artery bypass grafting  +  reduction annuloplasty for functional ischemic mitral regurgitation predictably reduces mitral regurgitation and relieves symptoms. This treatment of moderate to severe mitral regurgitation is associated with improved indices of ventricular function, improved New York Heart Association class, and excellent freedom from recurrent mitral insufficiency. Although long-term prognosis remains guarded, this multicenter study delineates the intermediate-term benefits of such an approach

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Not AvailableHeat stress is one of the major problems in agriculturally important cereal crops, especially wheat. Here, we have constructed a subtracted cDNA library from the endosperm of HS-treated (42°C for 2 h) wheat cv. HD2985 by suppression subtractive hybridization (SSH). We identified ~550 recombinant clones ranging from 200 to 500 bp with an average size of 300 bp. Sanger's sequencing was performed with 205 positive clones to generate the differentially expressed sequence tags (ESTs). Most of the ESTs were observed to be localized on the long arm of chromosome 2A and associated with heat stress tolerance and metabolic pathways. Identified ESTs were BLAST search using Ensemble, TriFLD, and TIGR databases and the predicted CDS were translated and aligned with the protein sequences available in pfam and InterProScan 5 databases to predict the differentially expressed proteins (DEPs). We observed eight different types of post-translational modifications (PTMs) in the DEPs corresponds to the cloned ESTs-147 sites with phosphorylation, 21 sites with sumoylation, 237 with palmitoylation, 96 sites with S-nitrosylation, 3066 calpain cleavage sites, and 103 tyrosine nitration sites, predicted to sense the heat stress and regulate the expression of stress genes. Twelve DEPs were observed to have transmembrane helixes (TMH) in their structure, predicted to play the role of sensors of HS. Quantitative Real-Time PCR of randomly selected ESTs showed very high relative expression of HSP17 under HS; up-regulation was observed more in wheat cv. HD2985 (thermotolerant), as compared to HD2329 (thermosusceptible) during grain-filling. The abundance of transcripts was further validated through northern blot analysis. The ESTs and their corresponding DEPs can be used as molecular marker for screening or targeted precision breeding program. PTMs identified in the DEPs can be used to elucidate the thermotolerance mechanism of wheat—a novel step toward the development of “climate-smart” wheat.Not Availabl

SSH Analysis of Endosperm Transcripts and Characterization of Heat Stress Regulated Expressed Sequence Tags in Bread Wheat

Heat stress is one of the major problems in agriculturally important cereal crops, especially wheat. Here, we have constructed a subtracted cDNA library from the endosperm of HS-treated (42°C for 2 h) wheat cv. HD2985 by suppression subtractive hybridization (SSH). We identified ~550 recombinant clones ranging from 200 to 500 bp with an average size of 300 bp. Sanger’s sequencing was performed with 205 positive clones to generate the differentially expressed sequence tags (ESTs). Most of the ESTs were observed to be localized on the long arm of chromosome 2A and associated with heat stress tolerance and metabolic pathways. Identified ESTs were BLAST search using Ensemble, TriFLD and TIGR databases and the predicted CDS were translated and aligned with the protein sequences available in pfam and InterProScan 5 databases to predict the differentially expressed proteins (DEPs). We observed eight different types of post-translational modifications (PTMs) in the DEPs corresponds to the cloned ESTs—147 sites with phosphorylation, 21 sites with sumoylation, 237 with palmitoylation, 96 sites with S-nitrosylation, 3066 calpain cleavage sites, and 103 tyrosine nitration sites, predicted to sense the heat stress and regulate the expression of stress genes. Twelve DEPs were observed to have transmembrane helixes (TMH) in their structure, predicted to play the role of sensors of HS. Quantitative Real-Time PCR of randomly selected ESTs showed very high relative expression of HSP17 under HS; up-regulation was observed more in wheat cv. HD2985 (thermotolerant), as compared to HD2329 (thermosusceptible) during grain-filling. The abundance of transcripts was further validated through northern blot analysis. The ESTs and their corresponding DEPs can be used as molecular marker for screening or targeted precision breeding program. PTMs identified in the DEPs can be used to elucidate the thermotolerance mechanism of wheat – a novel step towards the development of ‘climate-smart’ wheat

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Not AvailableHeat stress is one of the major problems in agriculturally important cereal crops, especially wheat. Here, we have constructed a subtracted cDNA library from the endosperm of HS-treated (42◦C for 2 h) wheat cv. HD2985 by suppression subtractive hybridization (SSH). We identified ∼550 recombinant clones ranging from 200 to 500 bp with an average size of 300 bp. Sanger’s sequencing was performed with 205 positive clones to generate the differentially expressed sequence tags (ESTs). Most of the ESTs were observed to be localized on the long arm of chromosome 2A and associated with heat stress tolerance and metabolic pathways. Identified ESTs were BLAST search using Ensemble, TriFLD, and TIGR databases and the predicted CDS were translated and aligned with the protein sequences available in pfam and InterProScan 5 databases to predict the differentially expressed proteins (DEPs). We observed eight different types of post-translational modifications (PTMs) in the DEPs corresponds to the cloned ESTs-147 sites with phosphorylation, 21 sites with sumoylation, 237 with palmitoylation, 96 sites with S-nitrosylation, 3066 calpain cleavage sites, and 103 tyrosine nitration sites, predicted to sense the heat stress and regulate the expression of stress genes. Twelve DEPs were observed to have transmembrane helixes (TMH) in their structure, predicted to play the role of sensors of HS. Quantitative Real-Time PCR of randomly selected ESTs showed very high relative expression of HSP17 under HS; up-regulation was observed more in wheat cv. HD2985 (thermotolerant), as compared to HD2329 (thermosusceptible) during grain-filling. The abundance of transcripts was further validated through northern blot analysis. The ESTs and their corresponding DEPs can be used as molecular marker for screening or targeted precision breeding program. PTMs identified in the DEPs can be used to elucidate the thermotolerance mechanism of wheat—a novel step toward the development of “climate-smart” wheat.Not Availabl