Comparative transcriptome profiling of contrasting finger millet (Eleusine coracana (L.) Gaertn) genotypes under heat stress.

BACKGROUND: Eleusine coracana (L.) Gaertn is a crucial C4 species renowned for its stress robustness and nutritional significance. Because of its adaptability traits, finger millet (ragi) is a storehouse of critical genomic resources for crop improvement. However, more knowledge about this crop's molecular responses to heat stress needs to be gained. METHODS AND RESULTS: In the present study, a comparative RNA sequencing analysis was done in the leaf tissue of the finger millet, between the heat-sensitive (KJNS-46) and heat-tolerant (PES-110) cultivars of Ragi, in response to high temperatures. On average, each sample generated about 24 million reads. Interestingly, a comparison of transcriptomic profiling identified 684 transcripts which were significantly differentially expressed genes (DEGs) examined between the heat-stressed samples of both genotypes. The heat-induced change in the transcriptome was confirmed by qRT-PCR using a set of randomly selected genes. Pathway analysis and functional annotation analysis revealed the activation of various genes involved in response to stress specifically heat, oxidation-reduction process, water deprivation, and changes in heat shock protein (HSP) and transcription factors, calcium signaling, and kinase signaling. The basal regulatory genes, such as bZIP, were involved in response to heat stress, indicating that heat stress activates genes involved in housekeeping or related to basal regulatory processes. A substantial percentage of the DEGs belonged to proteins of unknown functions (PUFs), i.e., not yet characterized. CONCLUSION: These findings highlight the importance of candidate genes, such as HSPs and pathways that can confer tolerance towards heat stress in ragi. These results will provide valuable information to improve the heat tolerance in heat-susceptible agronomically important varieties of ragi and other crops.

Gender Differences in Perceived Stress among Neurosurgeons: A Cross-Sectional Study.

Purpose: Surgeons are subjected to enhanced levels of work-related stress and women are likely to face unique challenges due to sub-optimal representation. The present study was conceived with a primary objective to study the gender differences faced by surgeons and neurosurgeons in particular. The secondary objective was to assess the correlation between the various stress inducers and busters of normal working and daily life and the mental state of neurosurgeons. Methods: The study was a cross-sectional, multi-centric study in which a structured questionnaire was sent to neurosurgeons through various neurosurgical forums of the country. A total of 93 complete responses were included in the study. The questions were broadly divided into four themes, demographic working data, stress inducers, stress busters, and questions to reflect the mental state of the respondent. Results: There were 74 males and 19 females in the study, with an average age of 39.34 ± 8.57 years. Statistically significant gender differences were noted in rating the out-patient department (OPD) hours, lectures and seminars, performing and assisting surgeries, attending conferences and working in the intensive care unit (ICU), with women scoring lower than men for these activities. There were no significant gender differences in the mental state of the respondents. A strong correlation was found between scoring for performance and assisting of surgery and the mental state questions, and a strong negative correlation was noted between music, playing games, going to the gym, practicing yoga and the mental state questions, indicative of a protective effect. Conclusions: There was no significant gender differences in the perceived stress levels among neurosurgeons. Women appeared more comfortable with certain normal activities of working. A strong negative correlation was reported for activities such as music, playing games, yoga, and going to the gym, indicating a protective effect.

RNA-Seq of Cyst Nematode Infestation of Potato (Solanum tuberosum L.): A Comparative Transcriptome Analysis of Resistant and Susceptible Cultivars.

Potato (Solanum tuberosum L.) is an important food crop worldwide, and potato cyst nematodes (PCNs) are among the most serious pests. The identification of disease resistance genes and molecular markers for PCN infestation can aid in crop improvement research programs against PCN infestation. In the present study, we used high-throughput RNA sequencing to investigate the comprehensive resistance mechanisms induced by PCN infestation in the resistant cultivar Kufri Swarna and the susceptible cultivar Kufri Jyoti. PCN infestation induced 791 differentially expressed genes in resistant cultivar Kufri Swarna, comprising 438 upregulated and 353 downregulated genes. In susceptible cultivar Kufri Jyoti, 2225 differentially expressed genes were induced, comprising 1247 upregulated and 978 downregulated genes. We identified several disease resistance genes (KIN) and transcription factors (WRKY, HMG, and MYB) that were upregulated in resistant Kufri Swarna. The differentially expressed genes from several enriched KEGG pathways, including MAPK signaling, contributed to the disease resistance in Kufri Swarna. Functional network analysis showed that several cell wall biogenesis genes were induced in Kufri Swarna in response to infestation. This is the first study to identify underlying resistance mechanisms against PCN and host interaction in Indian potato varieties.

Achieving maximum efficiency of Mungbean yellow mosaic India virus infection in mungbean by agroinoculation.

Mungbean is one of the important food legumes in the Indian-sub-continent. Yellow mosaic disease, caused by Mungbean yellow mosaic virus and Mungbean yellow mosaic India virus (MYMIV) poses a severe threat to its production. Agroinoculation has been the most preferred way to test the function of genomic components of these viruses. However, the available inoculation methods are not as efficient as whitefly transmission, thereby limiting their usage for screening and biological studies. We hereby report an efficient and reproducible agroinoculation method for achieving maximum (100%) efficiency using tandem repeat infectious agro-constructs of DNA A and DNA B of MYMIV. The present study targeted wounding of various meristematic tissues of root, shoot, parts of germinating seeds and also non-meristematic tissue of stem to test the suitable tissue types for maximum infection. Among the various tissues selected for, the inoculation on the epicotyl region showed maximum infectivity. Further, to enhance the infectivity of MYMIV, different concentrations of acetosyringone, incubation time and Agrobacterium cell density were also standardized. The incubation of wounded sprouted seeds in 1.0 OD of agroculture containing repeat construct of MYMIV for 2-4 h without acetosyringone followed by sowing in soil showed maximum infection of MYMIV within 10-12 days on the first trifoliate leaf. This standardized method is reproducible and has potential to screen germplasm lines and will be useful in mungbean biological/virological studies and breeding programmes. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s13205-021-03088-w.

Molecular analysis, infectivity and host range of Tomato leaf curl Karnataka virus associated with Corchorus yellow vein mosaic betasatellite.

Severe leaf curl disease of tomato (ToLCD) was noticed recently in the central parts of India and is an emerging threat to the cultivation of tomato. The genomic components of the begomovirus isolate, DNA A and betasatellite associated with ToLCD were cloned by rolling circle amplification method and sequenced. The sequence analysis revealed that the DNA A (2766 nt) of this isolate had the nucleotide identity of >91% with other strains of Tomato leaf curl Karnataka virus (ToLCKV), hence this isolate is proposed as a strain of ToLCKV, named as ToLCKV-Raipur. Similarly, the betasatellite molecule (1355 nt) had the highest identity of 91.1% with Corchorus yellow vein mosaic betasatellite (CoYVMB) and named as CoYVMB-Raipur. The full-length dimerized clones of these two genomic components were agroinoculated on natural (tomato), experimental (Nicotiana benthamiana) hosts and other 20 plant species belong to six different families. The severe leaf curl symptoms appeared only in the hosts, N. benthamiana, and in tomato inoculated with ToLCKV-Raipur alone and ToLCKV-Raipur with CoYVMB-Raipur after 8 and 16-18 days inoculation, respectively. This isolate was also transmissible to healthy tomato plants by whitefly from the tomato plant agroinoculated with ToLCKV-Raipur alone and with CoYVMB-Raipur and produced symptoms within 14-16 days after inoculation. Interestingly, this isolate infects horse gram and chilli by whitefly transmission and both the hosts showed positive for DNA A alone but not for betasatellite. Quantification of the genomic components of this isolate with the agroinoculated N. benthamiana samples by qRT-PCR results showed that the quantity of ToLCKV-Raipur was enhanced by three-fold while inoculated with CoYVMB-Raipur compared to ToLCKV-Raipur alone inoculated plants. However, CoYVMB-Raipur did not enhance the levels of ToLCKV-Raipur in the agroinoculated tomato plants. This is the first evidence of the natural co-occurrence of ToLCKV with betasatellite, CoYVMB causing ToLCD.

Distinct morpho-physiological and biochemical features of arid and hyper-arid ecotypes of Ziziphus nummularia under drought suggest its higher tolerance compared with semi-arid ecotype.

Tree species in the arid and semi-arid regions use various strategies to combat drought stress. Ziziphus nummularia (Burm. f.) Wight et Arn., native to the Thar Desert in India, is highly drought-tolerant. To identify the most drought-tolerant ecotype of Z. nummularia, one ecotype each from semi-arid (Godhra, annual rainfall >750 mm), arid (Bikaner, 250-350 mm) and hyper-arid (Jaisalmer, <150 mm) regions was selected along with two other Ziziphus species, Ziziphus mauritiana Lamk. and Ziziphus rotundifolia Lamk., and screened for parameters contributing to drought tolerance. Among these, Z. nummularia (Jaisalmer) (CIAHZN-J) was the most drought - tolerant. The tolerance nature of CIAHZN-J was associated with increased membrane stability, root length and number, length of hairs and thorns, root dry/fresh weight ratio, seed germination (at -0.5 MPa), proline content (31-fold), catalase and sugar content (two- to three-fold). Apart from these characteristics, it also exhibited the longest duration to reach highest cumulative drought stress rating, maintained higher relative water content for a longer period of time with reduced leaf size, leaf rolling and falling of older leaves, and displayed sustained shoot growth during drought stress. To determine drought tolerance in Ziziphus, we developed a morphological symptom-based screening technique in this study. Additionally, transcriptome profiling of CIAHZN-J in response to drought revealed the up-regulation of genes involved in sugar metabolism and transport, abscisic acid biosynthesis, osmoregulation, reactive oxygen species homeostasis and maintaining water potential. Expression profiles and semi-quantitative reverse transcription PCR results further correlated with the physiological and biochemical mechanisms. In conclusion, CIAHZN-J is an excellent genetic stock for the identification of drought-responsive genes and can also be deployed in crop improvement programs for drought tolerance.

Characterization and molecular modeling of Inositol 1,3,4 tris phosphate 5/6 kinase-2 from Glycine max (L) Merr.: comprehending its evolutionary conservancy at functional level.

Soybean genome encodes a family of four inositol 1,3,4 trisphosphate 5/6 kinases which belong to the ATP-GRASP group of proteins. Inositol 1,3,4 trisphosphate kinase-2 (GmItpk2), catalyzing the ATP-dependent phosphorylation of Inositol 1,3,4 trisphosphate (IP3) to Inositol 1,3,4,5 tetra phosphate or Inositol 1,3,4,6 tetra phosphate, is a key enzyme diverting the flux of inositol phosphate pool towards phytate biosynthesis. Although considerable research on characterizing genes involved in phytate biosynthesis is accomplished at genomic and transcript level, characterization of the proteins is yet to be explored. In the present study, we report the isolation and expression of single copy Itpk2 (948 bp) from Glycine max cv Pusa-16 predicted to encode 315 amino acid protein with an isoelectric point of 5.9. Sequence analysis revealed that GmITPK2 shared highest similarity (80%) with Phaseolus vulgaris. The predicted 3D model confirmed 12 α helices and 14 ß barrel sheets with ATP-binding site close to ß sheet present towards the C-terminus of the protein molecule. Spatio-temporal transcript profiling signified GmItpk2 to be seed specific, with higher transcript levels in the early stage of seed development. The present study using various molecular and bio-computational tools could, therefore, help in improving our understanding of this key enzyme and prove to be a potential target towards generating low phytate trait in nutritionally rich crop like soybean.

Transcriptome dynamics provide insights into long-term salinity stress tolerance in Triticum aestivum cv. Kharchia Local.

Kharchia Local, a wheat (Triticum aestivum) cultivar, is native to the saline-sodic soils of Pali district, Rajasthan, India and well known for its salinity stress tolerance. In the present study, we performed transcriptome sequencing to compare genome wide differential expression pattern between flag leaves of salinity stressed (15 EC) and control plants at anthesis stage. The 63.9 million paired end raw reads were assembled into 74,106 unigenes, of which, 3197 unigenes were found to be differentially expressed. Functional annotation analysis revealed the upregulation of genes associated with various biological processes including signal transduction, phytohormones signaling, osmoregulation, flavonoid biosynthesis, ion transport and ROS homeostasis. Expression pattern of fourteen differentially expressed genes was validated using qRT-PCR and was found to be consistent with the results of the transcriptome sequencing. Present study is the primary report on transcriptome profiling of Kharchia Local flag leaf under long-term salinity stress at anthesis stage. In conclusion, the data generated in this study can improve our knowledge in understanding the molecular mechanism of salinity stress tolerance. It will also serve as a valuable genomic resource in wheat breeding programs.