Phytohormones Accumulation and Distribution in Shoots and Roots of Haploid,Diploid and Tetraploid Barley Seedlings Derived from Microspore Culture

Phytohormones play important roles in plant growth and development,and polyploids are thought to be an important method for plant breeding.However,the relationship between ploidy and phytohormone is still unclear.In this study,barley at three ploidy levels were produced by microspore culture.Therefore,we further analyzed the phytohormone content in the shoots and roots of the three kinds of barley materials to study the effect of ploidy on phytohormones accumulation and distribution.The results showed that Abscisic acid(ABA),gibberellin(GA),jasmonic acid(JA),auxin(IAA),salicylic acid(SA)and cytokinin(CTK)were successfully determined in shoots and roots using LC-MS(liquid chromatography mass spectrometry).By comparing the shoots of the haploid and diploid plants,it was found that the distribution trend of the six phytohormones was consistent,and another consistent trend was found in the roots of the diploid and tetraploid plants.In addition,we further analyzed the shoot/root ratio of the different phytohormones to identify the potential differences for haploid,diploid and tetraploid.Here,the relationship between ploidy and phytohormone we provided would provide new insights into understanding the new phenotypes that occur in polyploid species.

Genome-Wide Identification of Candidate Genes Associated with β-glucan Traits in a Hulled and Hulless Barley (Hordeum vulgare L.) Population

Barley grain is a valuable source ofβ-glucan,which is an important component of dietary fiber with significant human health benefits.Although the genetic basis ofβ-glucan biosynthesis has been widely studied,a genome-wide association study(GWAS)is still required for a scan of the candidate genes related to the complex quantitative trait based on the high-quality barley reference genome.In this study,a GWAS was conducted using a population composed of 87 barley landraces(39 hulled and 48 hulless,β-glucan from 2.07%to 6.56%)with 191,098 nucleotide polymorphisms(SNPs)markers to cover the chromosomes with the highest density.The population was divided into four sub-populations(POP1~POP4),and theβ-glucan content in POP2 was significantly higher than that in other groups,in which most of the hulless barley landraces are from Qinghai-Tibet Plateau in China.Among seven SNP markers identified by GWAS,two(SNP2 and SNP3)of them showed positive correlation toβ-glucan trait and the remaining five(SNP1,SNP4,SNP5,SNP6 and SNP7)showed the negative relationship.Two candidate genes linked to SNP7,HORVU7Hr1G000320 and HORVU7Hr1G000040,belong to the nucleotide triphosphate hydrolase super-family which is probable to affect the activities ofβ-glucan synthase.Another candidate gene associated with SNP1,HORVU1Hr1G000010,is possibly involved in sugar response.In conclusion,our results provide new insights into the genetic basis ofβ-glucan accumulation in barley grains,and the discovery of new SNP markers distributed in each chromosome and the associated candidate genes will be valuable for the breeding of functional barley varieties with high β-glucan.

Transcriptome Profiling of Barley Cultivar Hua 30 MDEC in Response to Agrobacterium Infection

Agrobacterium-mediated transformation has been widely used in plants.However,the mechanism in plant cells’response to Agrobacterium infection was very complex.The mechanism of the determinants in host cell remains obscure,especially in barley,which is recalcitrant for Agrobacterium-mediated transformation.In the present study,microspore-derived embryogenic calli(MDEC)from barley elite cultivar were employed as unique subjects to characterize the mechanisms during the Agrobacterium infection process.Hua 30 MDEC can be successfully infected by Agrobacterium.RNA-sequencing at different infection points(0,2,6,12,24 hpi)was performed.The average expressional intensity of the whole genomics increased from 0 to 2 hpi,and then decreased subsequently.More upregulated than downregulated differentially expressed genes(DEGs)were counted at the same time.GO enrichment analysis showed that protein modification was significantly overrepresented in upregulated DEGs.Chromosome-related biological processes,gene expression and cellular metabolic processes were significantly overrepresented in downregulated DEGs.KEGG analysis showed that plant defense responses,phenylpropanoid biosynthesis and biosynthesis of amino acids were significantly enriched across the infection time course.Nine DEGs related to defense responses were identified.All DEGs were upregulated from 2 to 24 hpi.We speculate that these genes are possibly related to Agrobacterium infection.These findings will provide deep insights into the molecular events occurring during the process of Agrobacterium-mediated transformation.

Generating homozygous mutant populations of barley microspores by ethyl methanesulfonate treatment

Induced mutations are important for genetic research and breeding.Mutations induced by physical or chemical mutagenesis are usually heterozygous during the early generations.However,mutations must be fixed prior to phenotyping or field trials,which requires additional rounds of self-pollination.Microspore culture is an effective method to produce double-haploid(DH)plants that are fixed homozygotes.In this study,we conducted ethyl methanesulfonate(EMS)-induced mutagenesis of microspore cultures of barley(Hordeum vulgare)cultivar‘Hua30’and landrace‘HTX’.The EMS concentrations were negatively correlated with the efficiency of callus induction and the frequency of mutant plant regeneration.The two genotypes showed different regeneration efficiencies.The phenotypic variation of the regenerated M1 plants and the presence of genome-wide nucleotide mutations,revealed by whole-genome sequencing,highlight the utility of EMS-induced mutagenesis of isolated microspore cultures for developing DH mutants.Genome-wide analysis of the mutation frequency in the regenerated plants revealed that a considerable proportion of mutations resulted from microspore culture(somaclonal variation)rather than EMS-induced mutagenesis.In addition to producing a population of 1972 homozygous mutant lines that are available for future field trials,this study lays the foundation for optimizing the regeneration efficiency of DH plants and the richness of mutations(mainly by fine-tuning the mutagen dosage).