Alfalfa is the most widely cultivated forage legume, with approximately 30 million hectares planted worldwide. Genetic improvements in alfalfa have been highly successful in developing cultivars with exceptional winter hardiness and disease resistance traits. However, genetic improvements have been limited for complex economically important traits such as biomass. One of the major bottlenecks is the labor-intensive phenotyping burden for biomass selection. In this study, we employed two alfalfa fields to pave a path to overcome the challenge by using UAV images with fully automatic field plot segmentation for high-throughput phenotyping. The first field was used to develop the prediction model and the second field to validate the predictions. The first and second fields had 808 and 1025 plots, respectively. The first field had three harvests with biomass measured in May, July, and September of 2019. The second had one harvest with biomass measured in September of 2019. These two fields were imaged one day before harvesting with a DJI Phantom 4 pro UAV carrying an additional Sentera multispectral camera. Alfalfa plot images were extracted by GRID software to quantify vegetative area based on the Normalized Difference Vegetation Index. The prediction model developed from the first field explained 50–70% (R Square) of biomass variation in the second field by incorporating four features from UAV images: vegetative area, plant height, Normalized Green–Red Difference Index, and Normalized Difference Red Edge Index. This result suggests that UAV-based, high-throughput phenotyping could be used to improve the efficiency of the biomass selection process in alfalfa breeding programs.

Maize is the staple food crop in hilly areas of Nepal. Drought stress is one of the most important crop growth limiting factors leading to lower crop productivity in these areas. Maize seedlings are susceptible to drought stress. Nine lines (Arun-2, NML-1, Rampur Composite, RL-100, RL-105, Rl-106, RL-197, RML-18 and RML-8) of maize obtained from National Maize Research Program, Rampur, Chitwan, Nepal to study their tolerance to drought stress. An experiment was designed to study their drought tolerance by subjecting to four levels of osmotic potential (0,-5,-10 and-15 bar) induced by polyethylene glycol (PEG) 6000 in two factor completely randomized design with three replication at Biotechnology Division, National Agricultural Research Institute, Khumaltar, Lalitpur, Nepal. Effects of different levels of PEG 6000 were found significant on all traits studied viz. Germination rate (%), Speed of germination, Root length, Shoot length, Root Shoot ratio, Seedling length, Seedling fresh weight, Seedling dry weight and Vigor index. Line Arun-2 was recorded as best performer followed by Rampur Composite and RL-105 for all the traits analyzed. Lines RL-100, RL-106, RL-197, RML-18 and RML-8 showed poor performance under the same levels of PEG induced drought stress. Furthermore, the results also showed that maize manifests better genetic expression such as drought tolerance under severe moisture stress conditions in soil. Therefore, Lines Arun-2, Rampur Composite and RL-105 are suggested for varietal improvement program for drought stress conditions.

Alfalfa (Medicago sativa L.) is a perennial, outcrossing legume crop predominantly grown for hay, silage, or pasture. Intensive selection has resulted in dramatic improvement in fitness traits, including winter survival and disease resistance. However, there has been minimal improvement in other economically important traits, such as hay yield, which is still comparable to 30 years ago. Intensive phenotyping costs on this type of trait hinder high selection pressure to identify superior outcross individuals. Severe inbreeding depression inhibits the development of inbred lines with accumulated favorable alleles that exhibit heterosis. This review highlights the outcomes of inbreeding depression as well as the causes, including unmasking deleterious alleles and triggering self-incompatibility. We tracked the research efforts that unveil the genetic bases underlying deleterious alleles and self-incompatibility. The magnitudes of inbreeding depression were compared with the rate of heterozygous halved time in diploid and tetraploid organisms. To fill in the gaps between the controversy and existing hypotheses, we theorized a dosage dominant model of inheritance. The dosage dominant model is similar to the Mendelian dominance model, in which a genotype exhibits a dominant phenotype if there is a dominant allele (alphabet dominant). The difference is that in the dosage dominant model, a genotype will result in a dominant phenotype if the number of dominant alleles is equal to or greater than the number of recessive alleles. This review also includes a discussion on the development of pseudo inbreds and a hypothesis to identify deleterious alleles using bulked segregant analysis and consequently to purge deleterious alleles using marker-assisted selection, to progress toward the successful development of pure inbred lines in alfalfa.

Alfalfa (Medicago sativa L.) is a perennial, outcrossing legume crop predominantly grown for hay, silage, or pasture. Intensive selection has resulted in dramatic improvement in fitness traits, including winter survival and disease resistance. However, there has been minimal improvement in other economically important traits, such as hay yield, which is still comparable to 30 years ago. Intensive phenotyping costs on this type of trait hinder high selection pressure to identify superior outcross individuals. Severe inbreeding depression inhibits the development of inbred lines with accumulated favorable alleles that exhibit heterosis. This review highlights the outcomes of inbreeding depression as well as the causes, including unmasking deleterious alleles and triggering self-incompatibility. We tracked the research efforts that unveil the genetic bases underlying deleterious alleles and self-incompatibility. The magnitudes of inbreeding depression were compared with the rate of heterozygous halved time in diploid and tetraploid organisms. To fill in the gaps between the controversy and existing hypotheses, we theorized a dosage dominant model of inheritance. The dosage dominant model is similar to the Mendelian dominance model, in which a genotype exhibits a dominant phenotype if there is a dominant allele (alphabet dominant). The difference is that in the dosage dominant model, a genotype will result in a dominant phenotype if the number of dominant alleles is equal to or greater than the number of recessive alleles. This review also includes a discussion on the development of pseudo inbreds and a hypothesis to identify deleterious alleles using bulked segregant analysis and consequently to purge deleterious alleles using marker-assisted selection, to progress toward the successful development of pure inbred lines in alfalfa.

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Barley diseases are the major yield limiting factors for barley cultivation in Nepal. Stripe/Yellow rust (P. striformis f.sp. hordei and P. striformis f.sp. tritici), leaf rust (Puccinia hordei), and crown rust (P. coronata) are the major rust diseases in Nepal. Pyramiding resistance genes against all these rust diseases are possible through molecular marker assisted breeding. Sweden originated barley variety ‘Bonus’ is found resistant to stripe rust and having linked microsatellite markers for stripe rust and crown rust resistance. Similarly, Nepalese hull-less barley variety ‘Solu Uwa’ and Nepalese awn-less barley landrace NPGR Acc# 2478 have linked microsatellite markers for leaf rust resistance. Therefore, one polymorphic sequence tagged sites (STS) marker (ABG054) for stripe rust resistance, two polymorphic simple sequence repeats (SSR) markers (Bmac0144h and HVM049) for leaf rust and one polymorphic SSR marker (Bmag0006) for crown rust resistance were used to select the advanced barley lines (at F8 stage) from above parents. Field screening of stripe rust resistance was also conducted. Among 51 advanced and field disease resistance lines from Bonus/Solu Uwa cross, we have selected 10 pyramided lines for all three types of barley rust resistance. Similarly, among 39 advanced and field disease resistance lines from Bonus/NPGR Acc#2478 cross we have selected three pyramided lines and advanced for further yield testing for general cultivation purpose. The chances of losing the desired gene are high in late generation selection using molecular marker assisted selection (MAS); but the chances of getting agronomically superior varietal output will also increase.

Alfalfa (Medicago sativa L.) is the most cultivated forage legume around the world. Under a variety of growing conditions, forage yield in alfalfa is stymied by biotic and abiotic stresses including heat, salt, drought, and disease. Given the sessile nature of plants, they use strategies such as differential gene expression to respond to environmental cues. Transcription factors control the expression of genes that contribute to or enable tolerance and survival during periods of stress. Basic-leucine zipper (bZIP) transcription factors have been demonstrated to play a critical role in regulating plant growth and development as well as mediate the responses to abiotic stress in several species, including Arabidopsis thaliana, Oryza sativa, Lotus japonicus, and Medicago truncatula. However, there is little information about bZIP transcription factors in cultivated alfalfa. In the present study, 237 bZIP genes were identified in alfalfa from publicly available sequencing data. Multiple sequence alignments showed the presence of intact bZIP motifs in the identified sequences. Based on previous phylogenetic analyses in Arabidopsis thaliana, alfalfa bZIPs were similarly divided and fell into 10 groups. The physicochemical properties, motif analysis, and phylogenetic study of the alfalfa bZIPs revealed high specificity within groups. The differential expression of alfalfa bZIPs in a suite of tissues indicates that particular bZIP genes are specifically expressed at different developmental stages in alfalfa. Similarly, expression analysis in response to ABA, cold, drought, and salt stresses, indicates that a subset of bZIP genes are also differentially expressed and likely play a role in abiotic stress signaling and/or tolerance. These expression patterns were further verified by qRT-PCR. However, further functional characterization of bZIP transcription factors in alfalfa will help illuminate the role they play in stress tolerance mechanisms in legumes and facilitate the molecular breeding of stress tolerance in alfalfa.

Background Alfalfa (Medicago sativa L.) is the most cultivated forage legume around the world. Under a variety of growing conditions, forage yield in alfalfa is stymied by biotic and abiotic stresses including heat, salt, drought, and disease. Given the sessile nature of plants, they use strategies such as differential gene expression to respond to environmental cues. Transcription factors control the expression of genes that contribute to or enable tolerance and survival during periods of stress. Basic-leucine zipper (bZIP) transcription factors have been demonstrated to play a critical role in regulating plant growth and development as well as mediate the responses to abiotic stress in several species, including Arabidopsis thaliana, Oryza sativa, Lotus japonicus and Medicago truncatula. However, there is little information about bZIP transcription factors in cultivated alfalfa. ResultIn the present study, 237 bZIP genes were identified in alfalfa from publicly available sequencing data. Multiple sequence alignments showed the presence of intact bZIP motifs in the identified sequences. Based on previous phylogenetic analyses in Arabidopsis thaliana, alfalfa bZIPs were similarly divided and fell into 10 groups. The physico-chemical properties, motif analysis and phylogenetic study of the alfalfa bZIPs revealed high specificity within groups. The differential expression of alfalfa bZIPs in a suite of tissues indicates that bZIP genes are specifically expressed at different developmental stages in alfalfa. Similarly, expression analysis in response to ABA, cold, drought and salt stresses, indicates that a subset of bZIP genes are also differentially expressed and likely play a role in abiotic stress signaling and/or tolerance.ConclusionsTaken together, this work provides a framework for the future study of bZIPs in alfalfa and presents candidate bZIPs involved in stress-response signaling.