Xiaoli Jin

Evaluation of salinity tolerance and analysis of allelic function of HvHKT1 and HvHKT2 in Tibetan wild barley

Tibetan wild barley is rich in genetic diversity with potential allelic variation useful for salinity-tolerant improvement of the crop. The objectives of this study were to evaluate salinity tolerance and analysis of the allelic function of HvHKT1 and HvHKT2 in Tibetan wild barley. Salinity tolerance of 189 Tibetan wild barley accessions was evaluated in terms of reduced dry biomass under salinity stress. In addition, Na+ and K+ concentrations of 48 representative accessions differing in salinity tolerance were determined. Furthermore, the allelic and functional diversity of HvHKT1 and HvHKT2 was determined by association analysis as well as gene expression assay. There was a wide variation among wild barley genotypes in salt tolerance, with some accessions being higher in tolerance than cultivated barley CM 72, and salinity tolerance was significantly associated with K+/Na+ ratio. Association analysis revealed that HvHKT1 and HvHKT2 mainly control Na+ and K+ transporting under salinity stress, respectively, which was validated by further analysis of gene expression. The present results indicated that Tibetan wild barley offers elite alleles of HvHKT1 and HvHKT2 conferring salinity tolerance.

Genetic variation of HvCBF genes and their association with salinity tolerance in Tibetan annual wild barley.

The evaluation of both the genetic variation and the identification of salinity tolerant accessions of Tibetan annual wild barley (hereafter referred to as Tibetan barley) (Hordeum vulgare L. ssp. Spontaneum and H. vulgare L. ssp. agriocrithum) are essential for discovering and exploiting novel alleles involved in salinity tolerance. In this study, we examined tissue dry biomass and the Na+ and K+ contents of 188 Tibetan barley accessions in response to salt stress. We investigated the genetic variation of transcription factors HvCBF1, HvCBF3 and HvCBF4 within these accessions, conducting association analysis between these three genes and the respective genotypic salt tolerance. Salt stress significantly reduced shoot and root dry weight by 27.6% to 73.1% in the Tibetan barley lines. HvCBF1, HvCBF3 and HvCBF4 showed diverse sequence variation in amplicon as evident by the identification of single nucleotide polymorphisms (SNPs) and 3, 8 and 13 haplotypes, respectively. Furthermore, the decay of Linkage disequilibrium (LD) of chromosome 5 was 8.9 cM (r2<0.1). Marker bpb-4891 and haplotype 13 (Ps 610) of the HvCBF4 gene were significantly (P<0.05) and highly significantly (P<0.001) associated with salt tolerance. However, HvCBF1 and HvCBF3 genes were not associated with salinity tolerance. The accessions from haplotype 13 of the HvCBF4 gene showed high salinity tolerance, maintaining significantly lower Na+/K+ ratios and higher dry weight. It is thus proposed that these Tibetan barley accessions could be of value for enhancing salinity tolerance in cultivated barley.

Grain protein content variation and its association analysis in barley

BackgroundGrain protein content (GPC) is an important quality determinant for barley used as malt, feed as well as food. It is controlled by a complex genetic system. GPC differs greatly among barley genotypes and is also variable across different environments. It is imperative to understand the genetic control of barley GPC and identify the genotypes with less variation under the different environments.ResultsIn this study, 59 cultivated and 99 Tibetan wild barley genotypes were used for a genome-wide association study (GWAS) and a multi-platform candidate gene-based association analysis, in order to identify the molecular markers associated with GPC. Tibetan wild barley had higher GPC than cultivated barley. The significant correlation between GPC and diastatic power (DP), and malt extract confirmed the importance of GPC in determining malt quality. Diversity arrays technology (DArT) markers associated with barley GPC were detected by GWAS. In addition, GWAS revealed two HvNAM genes as the candidate genes controlling GPC. No association was detected between HvNAM1 polymorphism and GPC, while a single nucleotide polymorphism (SNP) (798, P < 0.01), located within the second intron of HvNAM2, was associated with GPC. There was a significant correlation between haplotypes of HvNAM1, HvNAM2 and GPC in barley.ConclusionsThe GWAS and candidate gene based-association study may be effectively used to determine the genetic variation of GPC in barley. The DArT markers and the polymorphism of HvNAM genes identified in this study are useful in developing high quality barley cultivars in the future. HvNAM genes could play a role in controlling barley GPC.

Determination of grain protein content by near-infrared spectrometry and multivariate calibration in barley

Grain protein content (GPC) is an important quality determinant in barley. This research aimed to explore the relationship between GPC and diffuse reflectance spectra in barley. The results indicate that normalizing, and taking first-order derivatives can improve the class models by enhancing signal-to-noise ratio, reducing baseline and background shifts. The most accurate and stable models were obtained with derivative spectra for GPC. Three multivariate calibrations including least squares support vector machine regression (LSSVR), partial least squares (PLS), and radial basis function (RBF) neural network were adopted for development of GPC determination models. The Lin_LSSVR and RBF_LSSVR models showed higher accuracy than PLS and RBF_NN models. Thirteen spectral wavelengths were found to possess large spectrum variation and show high contribution to calibration models. From the present study, the calibration models of GPC in barley were successfully developed and could be applied to quality control in malting, feed processing, and breeding selection.

A novel tissue-specific plantain β-1,3-glucanase gene that is regulated in response to infection by Fusarium oxysporum fsp. cubense

A new full-length β-1,3-glucanase cDNA, MpGlu, was isolated from a plantain (Musa paradisica) by the rapid amplification of cDNA ends (RACE) technique. Recombinant GST-MpGlu protein, expressed in E. coli, hydrolyzed (1→3),(1→6)-β-glucan of Laminaria digitata and inhibited the growth of Fusarium oxysporum fsp. cubense (race 4) suggesting that it is a β-1,3-glucanase. Southern blot analysis indicated that there is one copy of MpGlu in the plantain genome. MpGlu gene expression was detected in plantain leaves, peel, and pulp by RT-PCR. Northern blot analysis revealed that the expression of MpGlu was up-regulated by Fusarium infection. Subcellular localization analysis indicated that 28 residues at the N-terminal end are necessary for extracellular secretion, while 32 residues at the C-terminal end are necessary to target the protein into vacuoles.

The effect of H2O2 and abscisic acid (ABA) interaction on β-amylase activity under osmotic stress during grain development in barley

The effects of exogenous abscisic acid (ABA) and polyethylene glycol (PEG 6000) treatments on grain H(2)O(2), ABA and beta-amylase activity were studied during grain development in the spike culture experiments with variety Triumph and its ABA-insensitive mutant TL43 as the plant materials. The results showed that during grain development the two genotypes were similar in the pattern of ABA concentration change, but differed greatly in the pattern of H(2)O(2) concentration and beta-amylase activity changes. The beta-amylase activity was positively correlated with H(2)O(2) concentration, negatively correlated with ABA concentration, and it is mainly closely associated with continued high levels of ABA with respect to H(2)O(2). Water stress (PEG treatment) induced beta-amylase was associated with H(2)O(2) concentration but not with ABA concentration. Exogenous application of H(2)O(2) and Ascorbic acid (AsA) increased beta-amylase activity in Triumph but reduced that of TL43. However, the endogenous H(2)O(2) concentration in grains was always consistent with beta-amylase activity. A novel model was hypothesized from the current results to illustrate the relationship between H(2)O(2), ABA and beta-amylase synthesis for the barley exposed to abiotic stresses.

Determination of hemicellulose, cellulose and lignin content using visible and near infrared spectroscopy in Miscanthus sinensis

Lignocellulosic components including hemicellulose, cellulose and lignin are the three major components of plant cell walls, and their proportions in biomass crops, such as Miscanthus sinensis, greatly impact feed stock conversion to liquid fuels or bio-products. In this study, the feasibility of using visible and near infrared (VIS/NIR) spectroscopy to rapidly quantify hemicellulose, cellulose and lignin in M. sinensis was investigated. Initially, prediction models were established using partial least squares (PLS), least squares support vector machine regression (LSSVR), and radial basis function neural network (RBF_NN) based on whole wavelengths. Subsequently, 23, 25 and 27 characteristic wavelengths for hemicellulose, cellulose and lignin, respectively, were found to show significant contribution to calibration models. Three determination models were eventually built by PLS, LS-SVM and ANN based on the characteristic wavelengths. Calibration models for lignocellulosic components were successfully developed, and can now be applied to assessment of lignocellulose contents in M. sinensis.

Genotypic differences in callus induction and plant regeneration from mature embryos of barley (Hordeum vulgare L.)

An efficient induction system and regeneration protocol based on mature barley embryos were developed. Embryos isolated from mature seeds, dehusked by hand and inoculated with longitudinally bisected sections, showed low contamination and high primary callus-forming capability. The influences of nine culture media on primary callus induction and germination from the mature embryos of barley cultivars Golden Promise and Zaoshu 3 were analyzed. The results showed that the two cultivars had much higher values of primary callus induction in the B16M6D medium as compared to the other eight medium formulations, with a frequency of 74.3% and 78.4% for Golden Promise and Zaoshu 3, respectively. Furthermore, Zaoshu 3 demonstrated particularly high stability in callus induction over the different media, indicating its potential utilization in callus induction and regeneration for its good agronomic traits and wide adaption. There were significant differences amongst 11 barley genotypes in terms of primary callus induction in the optimum medium, with percentages of callus induction and germination response ranging from 17.9% to 78.4% and 2.8% to 47.4%, respectively. Green plantlets of Dong 17, Golden Promise, and Zaoshu 3 were successfully developed from primary calli through embryogenesis, with green plant differentiation frequencies ranging from 9.7% to 21.0% across genotypes.

Genetic Mapping of Quantitative Trait Loci Associated with β-amylase And Limit Dextrinase Activities and β-glucan and Protein Fraction Contents in Barley

High malting quality of barley (Hordeum vulgare L.) relies on many traits, such as β-amylase and limit dextrinase activities and β-glucan and protein fraction contents. In this study, interval mapping was utilized to detect quantitative trait loci (QTLs) affecting these malting quality parameters using a doubled haploid (DH) population from a cross of CM72 (six-rowed) by Gairdner (two-rowed) barley cultivars. A total of nine QTLs for eight traits were mapped to chromosomes 3H, 4H, 5H, and 7H. Five of the nine QTLs mapped to chromosome 3H, indicating a possible role of loci on chromosome 3H on malting quality. The phenotypic variation accounted by individual QTL ranged from 8.08% to 30.25%. The loci of QTLs for β-glucan and limit dextrinase were identified on chromosomes 4H and 5H, respectively. QTL for hordeins was coincident with the region of silica eluate (SE) protein on 3HS, while QTLs for albumins, globulins, and total protein exhibited overlapping. One locus on chromosome 3H was found to be related to β-amylase, and two loci on chromosomes 5H and 7H were found to be associated with glutelins. The identification of these novel QTLs controlling malting quality may be useful for marker-assisted selection in improving barley malting quality.

Association of HvLDI with limit dextrinase activity and malt quality in barley

Limit dextrinase (LD) is a unique de-branching enzyme involved in starch mobilization of barley grains during malting, and closely related to malt quality. Genotypic variation of LD activity is controlled by genetic factors and also affected by environmental conditions. Correlation analysis between LD activity and four malt quality parameters showed that LD activity was positively correlated with diastatic power, Kolbach index and the quality of malt extract, while negatively correlated with viscosity. The structure-based association analysis demonstrated that HvLDI, a gene encoding limit dextrinase inhibitor, was a major determinant of LD activity and malt quality. The single nucleotide polymorphisms associated with LD activity could be used in early generation selection for barley breeding.