Development of Novel KASP Markers for Improved Germination in Deep-Sown Direct Seeded Rice.

BACKGROUND: The lack of stable-high yielding and direct-seeded adapted varieties with better germination ability from deeper soil depth and availability of molecular markers are major limitation in achieving the maximum yield potential of rice under water and resource limited conditions. Development of high-throughput and trait-linked markers are of great interest in genomics-assisted breeding. The aim of present study was to develop and validate novel KASP (Kompetitive Allele-Specific PCR) markers associated with traits improving germination and seedling vigor of deep sown direct seeded rice (DSR). RESULTS: Out of 58 designed KASP assays, four KASP assays did not show any polymorphism in any of the eleven genetic backgrounds considered in the present study. The 54 polymorphic KASP assays were then validated for their robustness and reliability on the F1s plants developed from eight different crosses considered in the present study. The third next validation was carried out on 256 F3:F4 and 713 BC3F2:3 progenies. Finally, the reliability of the KASP assays was accessed on a set of random 50 samples from F3:F4 and 80-100 samples from BC3F2:3 progenies using the 10 random markers. From the 54 polymorphic KASP, based on the false positive rate, false negative rate, KASP utility in different genetic backgrounds and significant differences in the phenotypic values of the positive (desirable) and negative (undesirable) traits, a total of 12 KASP assays have been selected. These 12 KASP include 5 KASP on chromosome 3, 1 on chromosome 4, 3 on chromosome 7 and 3 on chromosome 8. The two SNPs lying in the exon regions of LOC_Os04g34290 and LOC_Os08g32100 led to non-synonymous mutations indicating a possible deleterious effect of the SNP variants on the protein structure. CONCLUSION: The present research work will provide trait-linked KASP assays, improved breeding material possessing favourable alleles and breeding material in form of expected pre-direct-seeded adapted rice varieties. The marker can be utilized in introgression program during pyramiding of valuable QTLs/genes providing adaptation to rice under DSR. The functional studies of the genes LOC_Os04g34290 and LOC_Os08g32100 possessing two validated SNPs may provide valuable information about these genes.

Integrating Association Mapping, Linkage Mapping, Fine Mapping with RNA Seq Conferring Seedling Vigor Improvement for Successful Crop Establishment in Deep Sown Direct-Seeded Rice.

BACKGROUND: Ongoing large-scale shift towards direct seeded rice (DSR) necessitates a convergence of breeding and genetic approaches for its sustenance and harnessing natural resources and environmental benefits. Improving seedling vigour remains key objective for breeders working with DSR. The present study aims to understand the genetic control of seedling vigour in deep sown DSR. Combined genome-wide association mapping, linkage mapping, fine mapping, RNA-sequencing to identify candidate genes and validation of putative candidate genes were performed in the present study. RESULTS: Significant phenotypic variations were observed among genotypes in both F3:4:5 and BC2F2:3 populations. The mesocotyl length showed significant positive correlation with %germination, root and shoot length. The 881 kb region on chromosome 7 reported to be associated with mesocotyl elongation. RNA-seq data and RT-PCR results identified and validated seven potential candidate genes. The four promising introgression lines free from linkage drag and with longer mesocotyl length, longer root length, semi-dwarf plant height have been identified. CONCLUSION: The study will provide rice breeders (1) the pre breeding material in the form of anticipated DSR adapted introgression lines possessing useful traits and alleles improving germination under deep sown DSR field conditions (2) the base for the studies involving functional characterization of candidate genes. The development and utilization of improved introgression lines and molecular markers may play an important role in genomics-assisted breeding (GAB) during the pyramiding of valuable genes providing adaptation to rice under DSR. Our results offer a robust and reliable package that can contribute towards enhancing genetic gains in direct seeded rice breeding programs.

Homeopathic Medicines in Second Wave of COVID-19: Prognostic Factor Research.

BACKGROUND: The clinical profile and course of COVID-19 evolved perilously in a second wave, leading to the use of various treatment modalities that included homeopathy. This prognostic factor research (PFR) study aimed to identify clinically useful homeopathic medicines in this second wave. METHODS: This was a retrospective, multi-centred observational study performed from March 2021 to May 2021 on confirmed COVID-19 cases who were either in home isolation or at COVID Care Centres in Delhi, India. The data were collected from integrated COVID Care Centres where homeopathic medicines were prescribed along with conventional treatment. Only those cases that met a set of selection criteria were considered for analysis. The likelihood ratio (LR) was calculated for the frequently occurring symptoms of the prescribed medicines. An LR of 1.3 or greater was considered meaningful. RESULTS: Out of 769 confirmed COVID-19 cases reported, 514 cases were selected for analysis, including 467 in home isolation. The most common complaints were cough, fever, myalgia, sore throat, loss of taste and/or smell, and anxiety. Most cases improved and there was no adverse reaction. Certain new symptoms, e.g., headache, dryness of mouth and conjunctivitis, were also seen. Thirty-nine medicines were prescribed, the most frequent being Bryonia alba followed by Arsenicum album, Pulsatilla nigricans, Belladonna, Gelsemium sempervirens, Hepar sulphuris, Phosphorus, Rhus toxicodendron and Mercurius solubilis. By calculating LR, the prescribing indications of these nine medicines were ascertained. CONCLUSION: Add-on use of homeopathic medicines has shown encouraging results in the second wave of COVID-19 in integrated care facilities. Further COVID-related research is required to be undertaken on the most commonly prescribed medicines.

Traumatic loss of total lower lip and chin: The anterolateral thigh flap provides an optimal solution.

BACKGROUND: Young female presented 15 days after traumatic avulsion of total lower lip and chin. Considering the functional and aesthetic needs of the lower lip and chin, it is indeed challenging for the reconstructive surgeon to provide aesthetically appealing lower facial subunit with maintenance of good oral seal and competence. The authors present this case of total lower lip and chin loss reconstructed as a single aesthetic unit with free anterolateral thigh flap followed by secondarily defining the aesthetic subunits into lower lip and chin by selective liposuction. METHOD: Free anterolateral thigh flap was harvested. The fascia lata, harvested separately, was divided to form slings over which distal part of the flap, designated to form the lower lip, was hung over and inset into the remnant gingivolabial sulcus mucosa. This folded distal part of the flap formed the lining and cover of the lower lip. RESULT: After resurfacing with free ALT flap, she was able to attain lip seal and resume oral feeds within 3-4 days. She presented six weeks after the first surgery with severely reduced mouth opening. This was due to excessively tight perioral slings. Slings were released and flap was selectively debulked using liposuction so as to define the lower lip and chin subunits. CONCLUSION: Free anterolateral thigh flap is an extremely versatile option in the armamentarium of free flaps available. Primary thinning and good colour match along with negligible donor site morbidity makes it meritorious over the radial forearm flap.

Convenient and ultra-sensitive fluorescence detection of bovine serum albumin by using Rhodamine-6G modified gold nanoparticles in biological samples.

This study comprises a convenient, rapid and very sensitive method for the determination of bovine serum albumin (BSA). The technique is based on fluorescence resonance energy transfer (FRET) between Rhodamine-6G (R6G) acting as donor and gold nanoparticles (AuNPs) acting as acceptors. This method takes advantage of AuNPs that have high quenching efficiency, therefore the absorption spectra range shifts from 521 to 635 nm when aggregation of the AuNPs takes place. Furthermore, when R6G was electrostatically self-adsorbed to the citrate-stabilized AuNPs surface the fluorescence intensity was quenched. After addition of BSA, the fluorescence intensity of the R6G recovered as BSA induced aggregation of the AuNPs and the adsorbed R6G was released to the solution. The recovery of intensity displays a linear relationship with BSA concentration over the range from 0.8 × 10-11  M to 5.6 × 10-11  M. The detection limit for BSA was found to be 4.58 × 10-11  M. The proposed method exhibited rapid analysis with high selectivity for BSA determination in human urine, blood and serum samples.

Prevalence, distribution and functional significance of the -237C to T polymorphism in the IL-12Rß2 promoter in Indian tuberculosis patients.

Cytokine/cytokine receptor gene polymorphisms related to structure/expression could impact immune response. Hence, the -237 polymorphic site in the 5' promoter region of the IL-12Rß2 (SNP ID: rs11810249) gene associated with the AP-4 transcription motif GAGCTG, was examined. Amplicons encompassing the polymorphism were generated from 46 pulmonary tuberculosis patients, 35 family contacts and 28 miscellaneous volunteers and sequenced. The C allele predominated among patients, (93.4%, 43/46), and in all volunteers and contacts screened, but the T allele was exclusively limited to patients, (6.5%, 3/46). The functional impact of this polymorphism on transcriptional activity was assessed by Luciferase-reporter and electrophoretic mobility shift assays (EMSA). Luciferase-reporter assays showed a significant reduction in transcriptional efficiency with T compared to C allele. The reduction in transcriptional efficiency with the T allele construct (pGIL-12Rb2-T), in U-87MG, THP-1 and Jurkat cell lines, were 53, 37.6, and 49.8% respectively, compared to the C allele construct (pGIL-12Rb2-C). Similarly, densitometric analysis of the EMSA assay showed reduced binding of the AP-4 transcription factor, to T compared to the C nucleotide probe. Reduced mRNA expression in all patients (3/3) harboring the T allele was seen, whereas individuals with the C allele exhibited high mRNA expression (17/25; 68%, p = 0.05). These observations were in agreement with the in vitro assessment of the promoter activity by Luciferase-reporter and EMSA assays. The reduced expression of IL-12Rß2 transcripts in 8 patients despite having the C allele was attributed to the predominant over expression of the suppressors (IL-4 and GATA-3) and reduced expression of enhancers (IFN-α) of IL-12Rß2 transcripts. The 17 high IL-12Rß2 mRNA expressers had significantly elevated IFN-α mRNA levels compared to low expressers and volunteers. Notwithstanding the presence of high levels of IL-12Rß2 mRNA in these patients elevated IFN-α expression could modulate their immune responses to Mycobacterium tuberculosis.