Amino acid transporter (AAT) gene family in Tartary buckwheat (Fagopyrum tataricum L. Gaertn.): Characterization, expression analysis and functional prediction.

Tartary buckwheat (Fagopyrum tataricum L. Gaertn., TB) is an ancient minor crop and an important food source for humans to supplement nutrients such as flavonoids and essential amino acids. Amino acid transporters (AATs) play critical roles in plant growth and development through the transport of amino acids. In this study, 104 AATs were identified in TB genome and divided into 11 subfamilies by phylogenetic relationships. Tandem and segmental duplications promoted the expansion of FtAAT gene family, and the variations of gene sequence, protein structure and expression pattern were the main reasons for the functional differentiation of FtAATs. Based on RNA-seq and qRT-PCR, the expression patterns of FtAATs in different tissues and under different abiotic stresses were analyzed, and several candidate FtAATs that might affect grain development and response to abiotic stresses were identified, such as FtAAP12 and FtCAT7. Finally, combined with the previous studies, the expression patterns and phylogenetic relationships of AATs in multiple species, the functions of multiple high-confidence FtAAT genes were predicted, and the schematic diagram of FtAATs in TB was initially drawn. Overall, this work provided a framework for further functional analysis of FtAAT genes and important clues for the improvement of TB quality and stress resistance.

Genome-Wide Development of Polymorphic Microsatellite Markers and Association Analysis of Major Agronomic Traits in Core Germplasm Resources of Tartary Buckwheat.

Tartary buckwheat (TB; Fagopyrum tataricum Gaertn.) is an important multigrain crop and medicinal plant, but functional genomics and molecular breeding research in this species have been lacking for quite some time. Here, genome-wide screening was performed to develop simple sequence repeat (SSR) markers associated with six major agronomic traits and the rutin contents of 97 core germplasm resources. A total of 40,901 SSR loci were identified; they were uniformly distributed throughout the TB genome, with a mean distance of 11 kb between loci. Based on these loci, 8,089 pairs of SSR primers were designed, and 101 primer pairs for polymorphic SSR loci were used to genotype the 97 core germplasm resources. The polymorphic SSR loci showed high genetic variation in these core germplasm resources, with an average polymorphic information content (PIC) value of 0.48. In addition, multiple SSR markers, such as SXAU8002 [100-grain weight (HGW)] and SXAU8006 [stem diameter (SD)], were found to be associated with agronomic traits in the two environments. Finally, based on gene functional annotation and homology analysis, a candidate gene, FtPinG0007685500, that may affect the node number and SD of the main stem by participating in lignin synthesis was identified. This study reports the mining of genome-wide SSR loci and the development of markers in TB, which can be used for molecular characterization of the germplasm in its gene pool. In addition, the detected markers and candidate genes could be used for marker-assisted breeding and functional gene cloning in TB.

Elucidation of the Regulatory Network of Flavonoid Biosynthesis by Profiling the Metabolome and Transcriptome in Tartary Buckwheat.

The characteristics of flavonoid metabolism in different Tartary buckwheat (TB) tissues and the related gene regulation network are still unclear at present. One hundred forty-seven flavonoids were identified from six TB tissues using the ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method. The roadmap of the rutin synthesis pathway was revealed. Through transcriptomic analysis it was revealed that the differentially expressed genes (DEGs) are mainly enriched in the "Phenylpropanoid biosynthesis" pathway. Fifty-two DEGs involved in the "flavonol synthesis" pathway were identified. The weighted gene correlation network analysis revealed four co-expression network modules correlated with six flavonol metabolites. Eventually, 74 genes revealed from MEblue and MElightsteelblue modules were potentially related to flavonol synthesis. Of them, 7 MYB transcript factors had been verified to regulate flavonoid synthesis. Furthermore, overexpressed FtMYB31 enhanced the rutin content in vivo. The present findings provide a dynamic flavonoid metabolism profile and co-expression network related to rutin synthesis and are thus valuable in understanding the molecular mechanisms of rutin synthesis in TB.

Comparative transcriptomic analysis reveals the regulatory mechanism of the gibberellic acid pathway of Tartary buckwheat (Fagopyrum tataricum (L.) Gaertn.) dwarf mutants.

BACKGROUND: Tartary buckwheat is an important minor crop species with high nutritional and medicinal value and is widely planted worldwide. Cultivated Tartary buckwheat plants are tall and have hollow stems that lodge easily, which severely affects their yield and hinders the development of the Tartary buckwheat industry. METHODS: Heifeng No. 1 seeds were treated with ethylmethanesulfonate (EMS) to generate a mutant library. The dwarf mutant ftdm was selected from the mutagenized population, and the agronomic characteristics giving rise to the dwarf phenotype were evaluated. Ultra-fast liquid chromatography-electrospray ionization tandem mass spectrometry (UFLC-ESI-MS/MS) was performed to determine the factors underlying the different phenotypes between the wild-type (WT) and ftdm plants. In addition, RNA sequencing (RNA-seq) was performed via the HiSeq 2000 platform, and the resulting transcriptomic data were analysed to identify differentially expressed genes (DEGs). Single-nucleotide polymorphism (SNP) variant analysis revealed possible sites associated with dwarfism. The expression levels of the potential DEGs between the WT and ftdm mutant were then measured via qRT-PCR and fragments per kilobase of transcript per million mapped reads (FPKM). RESULT: The plant height (PH) of the ftdm mutant decreased to 42% of that of the WT, and compared with the WT, the mutant and had a higher breaking force (BF) and lower lodging index (LI). Lower GA4 and GA7 contents and higher contents of jasmonic acid (JA), salicylic acid (SA) and brassinolactone (BR) were detected in the stems of the ftdm mutant compared with the WT. Exogenous application of GAs could not revert the dwarfism of the ftdm mutant. On the basis of the transcriptomic analysis, 146 homozygous SNP loci were identified. In total, 12 DEGs with nonsynonymous mutations were ultimately identified, which were considered potential candidate genes related to the dwarf trait. When the sequences of eight genes whose expression was downregulated and four genes whose expression was upregulated were compared, SKIP14, an F-box protein whose sequence is 85% homologous to that of SLY1 in Arabidopsis, presented an amino acid change (from Ser to Asn) and was expressed at a lower level in the stems of the ftdm mutant compared with the WT. Hence, we speculated that this amino acid change in SKIP14 resulted in a disruption in GA signal transduction, indirectly decreasing the GA content and downregulating the expression of genes involved in GA biosynthesis or the GA response. Further studies are needed to determine the molecular basis underlying the dwarf phenotype of the ftdm mutant. CONCLUSION: We report a Tartary buckwheat EMS dwarf mutant, ftdm, suitable for high-density planting and commercial farming. A significant decrease in GA4 and GA7 levels was detected in the ftdm mutant, and 12 DEGs expressed in the stems of the ftdm mutant were selected as candidates of the dwarfing gene. One nonsynonymous mutation was detected in the SKIP14 gene in the ftdm mutant, and this gene had a lower transcript level compared with that in the WT.

The Tartary Buckwheat Genome Provides Insights into Rutin Biosynthesis and Abiotic Stress Tolerance.

Tartary buckwheat (Fagopyrum tataricum) is an important pseudocereal crop that is strongly adapted to growth in adverse environments. Its gluten-free grain contains complete proteins with a well-balanced composition of essential amino acids and is a rich source of beneficial phytochemicals that provide significant health benefits. Here, we report a high-quality, chromosome-scale Tartary buckwheat genome sequence of 489.3 Mb that is assembled by combining whole-genome shotgun sequencing of both Illumina short reads and single-molecule real-time long reads, sequence tags of a large DNA insert fosmid library, Hi-C sequencing data, and BioNano genome maps. We annotated 33 366 high-confidence protein-coding genes based on expression evidence. Comparisons of the intra-genome with the sugar beet genome revealed an independent whole-genome duplication that occurred in the buckwheat lineage after they diverged from the common ancestor, which was not shared with rosids or asterids. The reference genome facilitated the identification of many new genes predicted to be involved in rutin biosynthesis and regulation, aluminum stress resistance, and in drought and cold stress responses. Our data suggest that Tartary buckwheat's ability to tolerate high levels of abiotic stress is attributed to the expansion of several gene families involved in signal transduction, gene regulation, and membrane transport. The availability of these genomic resources will facilitate the discovery of agronomically and nutritionally important genes and genetic improvement of Tartary buckwheat.

Effects of fully open-air [CO2] elevation on leaf photosynthesis and ultrastructure of Isatis indigotica fort.

Traditional Chinese medicine relies heavily on herbs, yet there is no information on how these herb plants would respond to climate change. In order to gain insight into such response, we studied the effect of elevated [CO2] on Isatis indigotica Fort, one of the most popular Chinese herb plants. The changes in leaf photosynthesis, chlorophyll fluorescence, leaf ultrastructure and biomass yield in response to elevated [CO2] (550±19 µmol mol(-1)) were determined at the Free-Air Carbon dioxide Enrichment (FACE) experimental facility in North China. Photosynthetic ability of I. indigotica was improved under elevated [CO2]. Elevated [CO2] increased net photosynthetic rate (P N), water use efficiency (WUE) and maximum rate of electron transport (J max) of upper most fully-expended leaves, but not stomatal conductance (gs), transpiration ratio (Tr) and maximum velocity of carboxylation (V c,max). Elevated [CO2] significantly increased leaf intrinsic efficiency of PSII (Fv'/Fm') and quantum yield of PSII(ΦPS II ), but decreased leaf non-photochemical quenching (NPQ), and did not affect leaf proportion of open PSII reaction centers (qP) and maximum quantum efficiency of PSII (Fv/Fm). The structural chloroplast membrane, grana layer and stroma thylakoid membranes were intact under elevated [CO2], though more starch grains were accumulated within the chloroplasts than that of under ambient [CO2]. While the yield of I. indigotica was higher due to the improved photosynthesis under elevated [CO2], the content of adenosine, one of the functional ingredients in indigowoad root was not affected.