Tolerant Landrace Research Articles

Evolutionary conservation of salinity responsive miRNAs from Indian wheat (Triticum aestivum) landrace, Kharchia Local

Being temporal, majority of abiotic stresses induces responses in plants that needs to be regulated in energy efficient manner. These regulations are carried out by many regulatory molecules including microRNAs and transcription factors. Like protein coding genes, miRNAs are also conserved across the plant species to exhibit their conserved function during growth, development and response to biotic or abiotic stresses. The present study was carried out during 2020 and 2021 at National phytotron facility, ICAR-Indian Agricultural Research Institute, New Delhi. Control and salinity treated plants of Kharchia Local, a highly salt tolerant landrace of wheat (Triticum aestivum L.) from India, were grown in hydroponics and after sequencing and analysis of small RNA data, salinity responsive mature miRNA sequences from Kharchia Local were analyzed for their evolutionary relationship with sequences from the public databases. The phylogenetic study, sequence similarity (identity scores) and multiple sequence comparison was used for evolutionary conservation analysis. The study revealed that, miRNA sequences from Kharchia Local are diverse and did not group with the salinity responsive miRNAs from the database except miR1551. Interestingly, a total of 25 known or conserved miRNA families were identified as salinity responsive across the plant species. The miRNAs from Kharchia Local appears to play regulatory role in novel mechanisms of salinity tolerance.

Insights into the responses of Akabare chili landraces to drought, heat, and their combined stress during pre-flowering and fruiting stages

Drought, heat, and their combined stress have increasingly become common phenomena in horticulture, significantly reducing chili production worldwide. The current study aimed to phenotype Akabare chili landraces (Capsicum spp.) in climate chambers subjected to drought and heat treatments during their early generative stage, focusing on PSII efficacy (Fv/Fm), net photosynthetic rate (PN), stomatal conductance (gs), leaf cooling, and biomass production. Six landraces were examined under heat and control conditions at 40/32 °C for 4 days and at 30/22 °C under drought and control conditions followed by a 5-day recovery under control conditions (30/22 °C, irrigated). Two landraces with higher (>0.77) and two with lower (<0.763) Fv/Fm during the stress treatments were later evaluated in the field under 55-day-long drought stress at the fruiting stage. In both treatments, stress-tolerant landraces maintained high Fv/Fm, PN, and better leaf cooling leading to improved biomass compared to the sensitive landraces. Agro-morpho-physiological responses of the tolerant and sensitive landraces during the early generative stage echoed those during the fruiting stage in the field. A climate chamber experiment revealed a 13.9 % decrease in total biomass under heat stress, a further 21.5 % reduction under drought stress, and a substantial 38.7 % decline under combine stress. In field conditions, drought stress reduced total biomass by 28.1 % and total fruit dry weight by 26.2 %. Tolerant landraces showed higher Fv/Fm, demonstrated better wilting scores, displayed a higher chlorophyll content index (CCI), and accumulated more biomass. This study validated lab-based results through field trials and identified two landraces, C44 and DKT77, as potential stress-tolerant genotypes. It recommends Fv/Fm, PN, and CCI as physiological markers for the early detection of stress tolerance.

Assessment of salt tolerance in Algerian oasis wheat landraces: An examination of biochemical, physiological, and agronomical traits

Wheat landraces cultivated in the oases of Algeria are are know for their resistance to abiotic stresses as a result of the extreme environmental constraints of the Sahara. As such, these landraces represent valuable breeding material for improving abiotic stress tolerance in wheat. This study was conducted to evaluate salt tolerance of bread and durum wheat from the Algerian oases. Ten wheat landraces from the Algerian oases were grown under prolonged salinity stress (150 mM NaCl) in greenhouse conditions. Data were assessed for 19 physiological, biochemical, and agronomical traits. The wheat landraces exhibited considerable variation in their salinity stress tolerance. The membership function value of salt tolerance identified Oum RokbaElhamra, Khellouf and Zeghlou as the most tolerant landraces while Bourione was identified as sensitive. The salt-tolerant and moderately tolerant wheat landraces maintained stable yields under conditions of salinity stress. Regression models constructed from MFVS and salt tolerance coefficients showed that for bread wheat, amino acid content and grain yield accounted for most of the variation in MFVS, while for durum wheat, the number of grains per plant and Na+ content explained the majority of observed differences in MFVS. Correlation analysis showed that the MFVS was significantly associated with grain yield, selectivity between K+ and Na+, and plant height. The results confirm that Algerian oasis wheat landraces are a valuable source given their salt tolerance and could be utilized in breeding programs seeking to improve salinity stress resilience in wheat.

Evaluating the physiological responses and identifying stress tolerance of Akabare chili landraces to individual and combined drought and heat stresses.

Abstract. Akabare chili (Capsicum annuum) contributes to Nepalese rural livelihoods but suffers from low productivity due to various abiotic stresses including drought and heat. This study aimed to assess the physiological responses of Akabare chili landraces to heat and drought stress, individually and together, and to identify stress-tolerant genotypes in the early vegetative stage. Selected eight Akabare chili landraces and chili variety 'Jwala' were subjected to control (30/22 °C day/night) and heat stress (40/32 °C) conditions with irrigation, and drought stress (30/22 °C) and combined drought-heat stress conditions without irrigation for 7 days, followed by a 5-day recovery under control condition. Stress-tolerant landraces showed better performance compared to sensitive ones in terms of efficacy of PS II (Fv/Fm), transpiration rate (E), net photosynthetic rate (PN), stomatal conductance (gs), leaf temperature depression, water use efficiency (WUE) and the ratio of stomata pore area to stomata area under stress conditions, resulting in improved biomass. Although all genotypes performed statistically similar under control conditions, their responses Fv/Fm, PN, E, gs and WUE were significantly reduced under thermal stress, further reduced under drought stress, and severely declined under the combination of both. Total biomass exhibited a 57.48 % reduction due to combined stress, followed by drought (37.8 %) and heat (21.4 %) compared to the control. Among the landraces, C44 showed the most significant gain in biomass (35 %), followed by DKT77 (33.48 %), while the lowest gain percentage was observed for C64C and PPR77 during the recovery phase (29 %). The tolerant landraces also showed a higher percentage of leaf cooling, chlorophyll content and leaf relative water content with fewer stomata but broader openings of pores. The study identifies potential stress-tolerant Akabare chili landraces and discusses the stress-tolerant physiological mechanisms to develop resilient crop varieties in changing climates.

Identifying Drought Tolerant Finger Millet Landraces for the Hills of Nepal

Finger millet is in the fourth rank among the major cereals in Nepal after rice, maize and wheat. Since it is cultivated in rainfed conditions, drought is one of the vital production constraints for this crop. With the objective of enhancing the utilization of native landraces conserved ex situ in genebank, 300 finger millet accessions were evaluated under drought stress and normal conditions at Khumaltar (1,360 masl), Lalitpur, Nepal during 2017 and 2018. In both the years, genotypes were significantly different for the yield and other agronomic characters. Eight different drought tolerant indices namely drought resistance index, geometric mean productivity, harmonic mean, mean productivity, stress susceptibility index, tolerance, yield index and yield stability index for each genotype were calculated using grain yield data recorded from the stress (Yds) and non-stress (Yns) experiment. Majority of the landraces were grouped under moderately susceptible group in both the years followed by moderately tolerant and susceptible group based on all the drought tolerant indices (DTIs). Five landraces, namely NGRC4849, NGRC6487, NGRC04852, NGRC03491 and NGRC6490 with average grain yield of 4670, 3624, 3426, 3191 and 3132 kg/ha, respectively, were identified as stable drought tolerant landraces compared to the released varieties. These landraces could be the potential sources of drought tolerance for finger millet improvement program for the mid-hills of Nepal.

Parental Polymorphism between Samba Mahsuri and False Smut Tolerant Landraces using SSR and InDel Markers

In the present study, 868 SSR and 475 InDel markers were selected to study parental polymorphism of two different crosses between recipient parent Samba Mahsuri and false smut donor linesIC379047 and IC334233 at ICAR-Indian Institute of Rice Research, Rajendranagar, Hyderabad. Out of 1343 markers used for parental polymorphism, 100 markers were polymorphic between parental lines Samba Mahsuri and landrace IC379047 and the total polymorphism percentage recorded was 7.44%. Ninety-nine markers exhibited polymorphism between parental lines Samba Mahsuri and IC334233 and 7.37% of total polymorphism percentage was recorded between the parents. The identified polymorphic markers could be further used for QTL mapping studies in false smut resistance.

Unraveling the genetic potential of native rice (Oryza sativa L.) landraces for tolerance to early-stage submergence.

Direct-seeded rice (DSR) is a promising alternative to the traditional puddled rice system. It has become more popular among rice growers as a result of socioeconomic shifts and global climate change. Although DSR offers advantages, rice plants experience greater anaerobic stress at sowing from unpredicted rainfall. Rice is unique among cereals in its ability to germinate under anaerobiosis. The coleoptile of rice rapidly elongates above the water surface to obtain more oxygen and enhance vigorous seedling growth. A panel of 115 landraces and four check varieties were subjected to anaerobic stress with a water level of 10 cm for up to 15 days. The present study observed significant variation in anaerobic germination percentage (AGP) (10%-100%) and anaerobic vigor index (AVI) (150-4,433). Landraces Karuthakar, Poovan samba, Mattaikar, Edakkal, Manvilayan, and Varappu kudainchan were identified as genotypes tolerant to early water submergence. The shoot and root length of susceptible landraces were significantly lower than the tolerant landraces under hypoxia condition, implying that landraces with longer shoots and roots had a higher survival rate. The response index substantiated this. The results clearly show that tolerant and moderately tolerant landraces possessed higher mean values for root and shoot lengths than susceptible landraces. The landraces grouped under the long-bold category had superior AGP and AVI scores to other grain type groups. This raises the possibility that differences in kernel breadth, which is linked to grain type, could affect anaerobic germination potential. Molecular confirmation using gene-specific markers, viz., DFR, TTP_G4, RM478, RM208, and RM24161, for which the polymorphic information content (PIC) value ranged from 0.36 (RM478) to 0.68 (RM206) suggests that this diverse panel of landraces must be assessed further using advanced molecular tools to precisely clarify the genetic mechanism behind this phenomenon. The tolerant landraces thus identified may become donors in breeding programs. The introduction of these traits would contribute to the development of rice varieties tolerant to anaerobic stress, resulting in sustainable yields. This solution could promote the DSR system across the world.

English

Twenty-one Nigerian rice landraces were screened for tolerance to phosphate deficiency in nutrient medium using selected morphological indices from which Phosphate Deficiency Tolerance Index (PDTI) was evaluated. All landraces were analyzed for the presence of four phosphate uptake 1 (Pup-1) gene-linked markers while relative expression levels of two Pup-1 genes were evaluated in selected landraces under zero, normal and excess phosphate. PDTI analysis grouped the landraces into negative, low positive or high positive PDTI categories depending on whether performance at zero P was better than normal P, comparable to normal P, or less than normal P, respectively. However, irrespective of PDTI grouping, each landrace had at least one Pup-1 gene marker. Under zero P, Phillipine landrace showed no superior expression of OsPupK05-1 and OsPupK04-1 genes despite belonging to negative PDTI group. Thus, PDTI-based characterization of the landraces was not completely consonant with the presence or expression levels of the Pup-1 genes, suggesting a possible influence of other P deficiency tolerant genes. However, with a combination of negative PDTI, superior performance in root and shoot traits under zero P, and possession of at least 3 Pup 1 genes, Dantala Mass, Ankulyan, and Variety 44 may be regarded as P tolerant landraces. &nbsp; Key words: Oryza sativa, phosphate deficiency tolerance index, RT-PCR, Pup genes.

Salt tolerance QTLs of an endemic rice landrace, Horkuch at seedling and reproductive stages

Salinity has a significant negative impact on production of rice. To cope with the increased soil salinity due to climate change, we need to develop salt tolerant rice varieties that can maintain their high yield. Rice landraces indigenous to coastal Bangladesh can be a great resource to study the genetic basis of salt adaptation. In this study, we implemented a QTL analysis framework with a reciprocal mapping population developed from a salt tolerant landrace Horkuch and a high yielding rice variety IR29. Our aim was to detect genetic loci that contributes to the salt adaptive responses of the two different developmental stages of rice which are very sensitive to salinity stress. We identified 14 QTLs for 9 traits and found that most are unique to specific developmental stages. In addition, we detected a significant effect of the cytoplasmic genome on the QTL model for some traits such as leaf total potassium and filled grain weight. This underscores the importance of considering cytoplasm-nuclear interaction for breeding programs. Finally, we identified QTLs co-localization for multiple traits that highlights the possible constraint of multiple QTL selection for breeding programs due to different contributions of a donor allele for different traits.

Screening of rice germplasms (Oryza sativa L.) for seedling stage cold tolerance utilizing morphological and molecular markers

Cold temperature is a devastating abiotic stress for rice production, especially in the areas where low temperature prevails throughout the crop duration or in particular growth stages. Boro cultivation season in eastern India is such a replica of rice cultivation under occasional colder climate. Therefore, it is imperative to study a large collection of germplasms for potential cold tolerant stocks. In this study, 106 germplasms from eastern India were cultivated during Boro season in field as well as under 10 °C and 28 °C temperature conditions inside plant growth chamber. Six seedling growth parameters were used as phenotypic markers and 56 cold tolerance quantitative trait loci linked simple sequence repeats (SSRs) were utilized as molecular markers. Leaf yellowing and germination index were also incorporated within the framework of cold tolerance screening to enrich the multidimensional approach. Based on the k-means clustering for each morphological parameter in each condition, the genotypes were ranked and the resulting total ranks for each genotype eventually grouped them into four clusters, i.e. highly susceptible, susceptible, moderately tolerant and tolerant as different level of cold tolerance. Among the screened SSRs, only 33 were polymorphic and most informative were RM336, RM566 and RM1349 due to higher marker efficiency threshold. Combining both morphology and molecular analysis, highly tolerant landraces such as Neera, Barnamali, Khaiya Boro and genotypes such as Purbachi, CB1, Gotra, UPR 103-80 were identified which may be of greater use in development of high yielding cold tolerant genotypes.

Synergistic Modulation of Seed Metabolites and Enzymatic Antioxidants Tweaks Moisture Stress Tolerance in Non-Cultivated Traditional Rice Genotypes during Germination.

Traditional rice landraces are treasures for novel genes to develop climate-resilient cultivars. Seed viability and germination determine rice productivity under moisture stress. The present study evaluated 100 rice genotypes, including 85 traditional landraces and 15 improved cultivars from various agro-ecological zones of Tamil Nadu, along with moisture-stress-susceptible (IR 64) and moisture-stress-tolerant (IR 64 Drt1) checks. The landraces were screened over a range of osmotic potentials, namely (−) 1.0 MPa, (−) 1.25 MPa and (−) 1.5 MPa, for a period of 5 days in PEG-induced moisture stress. Physio-morphological traits, such as rate of germination, root and shoot length, vigor index, R/S ratio and relative water content (RWC), were assessed during early moisture stress at the maximum OP of (−) 1.5 MPa. The seed macromolecules, phytohormones (giberellic acid, auxin (IAA), cytokinin and abscisic acid), osmolytes and enzymatic antioxidants (catalase and superoxide dismutase) varied significantly between moisture stress and control treatments. The genotype Kuliyadichan registered more IAA and giberellic acid (44% and 35%, respectively, over moisture-stress-tolerant check (IR 64 Drt1), whereas all the landraces showed an elevated catalase activity, thus indicating that the tolerant landraces effectively eliminate oxidative damages. High-performance liquid chromatography analysis showed a reduction in cytokinin and an increase in ABA level under induced moisture stress. Hence, the inherent moisture-stress tolerance of six traditional landraces, such as Kuliyadichan, Rajalakshmi, Sahbhagi Dhan, Nootripathu, Chandaikar and Mallikar, was associated with metabolic responses, such as activation of hydrolytic enzymes, hormonal crosstalk, ROS signaling and antioxidant enzymes (especially catalase), when compared to the susceptible check, IR 64. Hence, these traditional rice landraces can serve as potential donors for introgression or pyramiding moisture-stress-tolerance traits toward developing climate-resilient rice cultivars.

Proto Kranz-like leaf traits and cellular ionic regulation are associated with salinity tolerance in a halophytic wild rice

Species of wild rice (Oryza spp.) possess a wide range of stress tolerance traits that can be potentially utilized in breeding climate-resilient cultivated rice cultivars (Oryza sativa) thereby aiding global food security. In this study, we conducted a greenhouse trial to evaluate the salinity tolerance of six wild rice species, one cultivated rice cultivar (IR64) and one landrace (Pokkali) using a range of electrophysiological, imaging, and whole-plant physiological techniques. Three wild species (O. latifolia, O. officinalis and O. coarctata) were found to possess superior salinity stress tolerance. The underlying mechanisms, however, were strikingly different. Na+ accumulation in leaves of O. latifolia, O. officinalis and O. coarctata were significantly higher than the tolerant landrace, Pokkali. Na+ accumulation in mesophyll cells was only observed in O. coarctata, suggesting that O. officinalis and O. latifolia avoid Na+ accumulation in mesophyll by allocating Na+ to other parts of the leaf. The finding also suggests that O. coarctata might be able to employ Na+ as osmolyte without affecting its growth. Further study of Na+ allocation in leaves will be helpful to understand the mechanisms of Na+ accumulation in these species. In addition, O. coarctata showed Proto Kranz-like leaf anatomy (enlarged bundle sheath cells and lower numbers of mesophyll cells), and higher expression of C4-related genes (e.g., NADPME, PPDK) and was a clear outlier with respect to salinity tolerance among the studied wild and cultivated Oryza species. The unique phylogenetic relationship of O. coarctata with C4 grasses suggests the potential of this species for breeding rice with high photosynthetic rate under salinity stress in the future.

Marker based genetic variability analysis of rice (Oryza sativa L.) landraces for drought tolerance

Under the present study, assessment of genetic variability of 60 numbers of rice landraces was carried out under morphological and molecular level for selection of promising drought tolerant genotypes under upland ecosystem of North East India. Predominance of additive gene action on characters like leaf rolling index, root: culm ratio, grain yield and other yield contributing traits like plant height came out as effective parameters for selection of drought stress tolerant landraces. Molecular analysis revealed moderate genetic diversity with average Polymorphic Information Content values of 0.44 across the rice landraces. No specific clustering patterns of landraces against some morphological traits numbers of productive tillers per plant, numbers of filled grains per plant etc were found. This signifies the polygenic nature of the quantitative characters and influence of environment on them. Precise correlation between the morphological performance of the landraces and their clustering pattern under molecular analysis was found to be effective in identifying suitable landraces like Chikanswarikabar as a promising parent for future breeding programme and also to formulate efficient strategies for sustainable management of rice landraces under rain fed ecosystem.

Proteome Mapping of South African Cassava Mosaic Virus-Infected Susceptible and Tolerant Landraces of Cassava.

The production of cassava is threatened by the geminivirus South African cassava mosaic virus (SACMV), which causes cassava mosaic disease. Cassava landrace TME3 shows tolerance to SACMV, while T200 is highly susceptible. This study aimed to identify the leaf proteome involved in anti-viral defence. Liquid chromatography mass spectrometry (LC-MS) identified 2682 (54 differentially expressed) and 2817 (206 differentially expressed) proteins in both landraces at systemic infection (32 days post infection) and symptom recovery (67 days post infection), respectively. Differences in the number of differentially expressed proteins (DEPs) between the two landraces were observed. Gene ontology analysis showed that defence-associated pathways such as the chloroplast, proteasome, and ribosome were overrepresented at 67 days post infection (dpi) in SACMV-tolerant TME3. At 67 dpi, a high percentage (56%) of over-expressed proteins were localized in the chloroplast in TME3 compared to T200 (31% under-expressed), proposing that chloroplast proteins play a role in tolerance in TME3. Ribosomal_L7Ae domain-containing protein (Manes.12G139100) was over-expressed uniquely in TME3 at 67 dpi and interacts with the ribosomal protein Sac52 (RPL10). RPL10 is a known key player in the NIK1-mediated effector triggered immunity (ETI) response to geminivirus infection, indicating a possible role for Sac52 in SACMV recovery in TME3. In conclusion, differential protein expression responses in TME3 and T200 may be key to unravel tolerance to CMD.

Screening of rice genotypes for salt tolerance by physiological and biochemical characters

Crop production is unexpectedly hampered by different abiotic stresses. Salinity is one of the leading stresses, which snappishly hampers plant developmental progression. Local rice landraces exhibit noticeable salt tolerance as well as high yield. However, research is scarce about the physio-biochemical responses of local landraces and modern cultivar under saline conditions. Therefore, the present experiment was designed to reveal the physio-biochemical responses of local landraces and modern cultivar under salinity stress. Five landraces (Jotai, Icheburogolghor, Morishal, Chapail, Kumro buro) and two modern cultivars (BR23 and BRRI dhan41) were subjected to 0, 20, 40, 60 and 80 mM NaCl treatment. The effects of salt stress on morphological parameters, proline contents, and activities of antioxidant enzymes were assessed. Salt stress reduces the morphological parameters of all tested cultivars. The Morishal and BRRI dhan41 exhibited higher growth of plant and physiological parameters than other cultivars under the highest salinity. The catalase (CAT) and ascorbate peroxidase (APX), exhibited a significant increase whereas peroxidase (POX) activity significantly declined in all the cultivars under salinity stress. Morishal and BRRI dhan41 showed the highest proline content under the maximum saline condition. These results suggest that the high tolerant landrace and modern cultivars were Morishal and BRRI dhan41 respectively. These results also suggest that Morishal and BRRI dhan41 exhibited high tolerance to salinity by enhancing proline content and antioxidant enzyme activities.

Genotypic variation in biochemical and physiological responses of fenugreek (Trigonella foenum-graecum L.) landraces to prolonged drought stress and subsequent rewatering

Sixteen fenugreek landraces from Iran and two landraces from Egypt and India were evaluated for their biochemical and physiological responses to prolonged drought stress and recovery. The landraces were classified into six and three groups during drought stress and recovery, respectively, by cluster analysis. More tolerant landraces showed higher stomatal conductance and transpiration rate. The high potential recovery of landraces after rewatering can be related to high levels of SOD activity and proline accumulation. Ardabil and Shushtar landraces were the most drought tolerant landraces, while Shiraz exhibited the highest potential for recovery after drought. Varamin and Mashhad landraces were the weakest landraces during drought stress and recovery. Some Iranian landraces showed better responses to drought stress and recovery than exotic landraces. Our results indicated that high sensitivity to drought stress does not necessarily mean low recovery potential in landraces. Besides, wide genotypic variation in biochemical and physiological traits among the landraces indicates the importance of these traits in selecting for tolerance to drought stress, especially in arid and semi-arid environments.

Biochemical performance of some Sub1 rice (Oryza sativa) genotypes under submergence stress in Odisha

Recurring floods in Asia cause poor crop establishment. Yields decline drastically when plants are completely submerged for a few days. Traditional rice genotypes predominate because they have acquired moderate tolerance to flooding but they carry the penalty of inherently lower grain yields. Genotypes with tolerance to complete submergence were recently developed in the background of popular genotypes by transferring the submergence tolerance gene SUBMERGENCE1 (Sub1) from the highly tolerant Indian landrace FR13A. The present study evaluated thirteen pairs of Sub1 near-isogenic lines (NILs) together with FR13A and other check genotypes in pot culture conditions to assess the survival and growth processes occurring during submergence and recovery that are associated with Sub1. The present experiment was conducted in Department of Plant physiology, OUAT, Bhubaneswar during kharif 2017 and kharif 2019 to screen out the NILs rice genotypes for submergence adaptation traits under coastal regions of Odisha. The present study indicated that among the twenty genotypes, IR-85086-Sub 33-3-2-1(4.87 t ha-1), IR-88760-Sub 93-3-3 (4.7 t ha-1) and Swarna Sub -1 (4.57 t ha-1) contributed highest yield under submergence.

Screening of tomato landraces for drought tolerance based on growth and chlorophyll fluorescence analyses

To investigate the effects of water withholding on 17 tomato (Solanum lycopersicum L.) landraces collected from different climatic zones of Iran and two commercial hybrids, the polyphasic OJIP fluorescence transient, relative water content (RWC), electrolyte leakage (EL) and vegetative growth parameters were analyzed. Duncan’s multiple range test (DMRT) for all the studied parameters and drought factor index (DFI) based on performance index on the absorption basis (PIabs) were used for screening the plants based on their tolerance to drought condition. Result showed that compared to the control plants, vegetative growth parameters, RWC, PIabs, relative maximal variable fluorescence (FM/F0), maximum quantum efficiency of PSII (FV/FM), quantum yield of electron transport (ΦE0) and electron transport flux per reaction center (RC) (ET0/RC) were decreased, whereas, EL, quantum yield of energy dissipation (ΦD0), specific energy fluxes per RC for energy absorption (ABS/RC) and dissipated energy flux (DI0/RC), which are closely related to the incidence of photoinhibition were increased in plants exposed to water withholding. DMRT and DFI screening results clearly categorized the landraces into three groups (tolerant, moderately sensitive and sensitive). Tolerant landraces showed less change for most of the measured parameters compared to sensitive and moderately sensitive landraces. We found that adapted landraces to dry climates had a higher tolerance to drought stress. Principal component analysis (PCA) revealed that FM/F0, FV/FM, ΦE0, ΦD0, PIabs, ABS/RC, ET0/RC and DI0/RC parameters are the most sensitive parameters for detection of impact of drought stress on tomato plants. In conclusion, the eight parameters have the potential to identify the drought injury in tomato seedlings.

A cassava protoplast system for screening genes associated with the response to South African cassava mosaic virus

BackgroundThe study of transient gene expression in cassava plants during virus infection using existing protocols is laborious and may take approximately fifteen weeks due to cassava’s recalcitrance to transformation. The combination of a protoplast system with CRISPR-mediated gene editing promises to shorten the turnaround time from plant tissue culture to high-throughput gene expression screening for candidate genes. Here, we detail a protocol for screening genes associated with the response to South African cassava mosaic virus (SACMV) in cassava protoplasts, with reference to the ubiquitin E3 ligase gene, MeE3L.MethodsCassava protoplasts of model, and SACMV-susceptible and -tolerant genotypes, were transformed with SACMV infectious clones and/or a CRISPR-editing construct targeting the MeE3L using PEG4000-mediated transfection. DNA and RNA were extracted from transformed protoplasts at 24 h post-transfection. Relative SACMV DNA accumulation was determined via qPCR using DpnI-digested total DNA, MeE3L relative expression was determined via reverse transcriptase qPCR, and results were analysed using one-way ANOVA, Tukey’s HSD test and the 2−ΔΔCTstatistical method. The MeE3L exonic region was sequenced on the ABI 3500XL Genetic Analyzer platform; and sequences were analysed for mutations using MAFTT and MEGA-X software. Construction of a phylogenetic tree was done using the Maximum Likelihood method and Jones-Taylor-Thornton (JTT) matrix-based model.ResultsThe differential expression of unedited and mutant MeE3L during SACMV infection of model, susceptible and tolerant cassava protoplasts was determined within 7 weeks after commencement of tissue culture. The study also revealed that SACMV DNA accumulation in cassava protoplasts is genotype-dependent and induces multiple mutations in the tolerant landrace MeE3L homolog. Notably, the susceptible cassava landrace encodes a RINGless MeE3Lwhich is silenced by SACMV-induced mutations. SACMV also induces mutations which silence the MeE3L RING domain in protoplasts from and tolerant cassava landraces.ConclusionsThis protocol presented here halves the turnaround time for high-throughput screening of genes associated with the host response to SACMV. It provides evidence that a cassava E3 ligase is associated with the response to SACMV and forms a basis for validation of these findings by in planta functional and interaction studies.

Marker-assisted identification of novel genetic lines for salinity tolerance and their categorization for utilization in development of hybrid rice (Oryza sativa L.)

Identification of new sources of salinity-tolerant genotypes is prerequisite for rice breeding programs in different saline ecosystems. In the present experiment, we characterized 177 landraces from the Western Ghats of Karnataka under natural saline field conditions for two years using morpho-physiological and grain quality parameters. Significant variation was present among landraces for seedling stage and reproductive stage salinity tolerance. The nutrient composition analysis of grain revealed an increase in average grain protein and carbohydrate content under saline conditions. Evaluation of twenty-two SSR markers associated with the Saltol region validated RM140, RM1287 and RM562 as best markers to classify landraces for saline tolerance. Polymorphism Information Content and genetic diversity indices showed that the markers RM10748 and RM10864 were highly useful for distinguishing landraces. Further, to benefit the exploitation of heterosis, eleven maintainers were identified among tolerant landraces and these genotypes could be further developed into male sterile lines for production of salinity-tolerant rice hybrids. Comparison with ‘Pokkali’ for morpho-physiological traits along with molecular confirmation showed that the landraces ‘Doddabaikalu,’ Kalaadikonda,’ Gajagunda’ and ‘Anekombina batha’ were superior donors carrying genomic regions for salinity tolerance.