Genome-wide analysis of filamentous temperature-sensitive H protease (ftsH) gene family in soybean.

BACKGROUND: The filamentous temperature-sensitive H protease (ftsH) gene family belongs to the ATP-dependent zinc metalloproteins, and ftsH genes play critical roles in plant chloroplast development and photosynthesis. RESULTS: In this study, we performed genome-wide identification and a systematic analysis of soybean ftsH genes. A total of 18 GmftsH genes were identified. The subcellular localization was predicted to be mainly in cell membranes and chloroplasts, and the gene structures, conserved motifs, evolutionary relationships, and expression patterns were comprehensively analyzed. Phylogenetic analysis of the ftsH gene family from soybean and various other species revealed six distinct clades, all of which showed a close relationship to Arabidopsis thaliana. The members of the GmftsH gene family were distributed on 13 soybean chromosomes, with intron numbers ranging from 3 to 15, 13 pairs of repetitive segment. The covariance between these genes and related genes in different species of Oryza sativa, Zea mays, and Arabidopsis thaliana was further investigated. The transcript expression data revealed that the genes of this family showed different expression patterns in three parts, the root, stem, and leaf, and most of the genes were highly expressed in the leaves of the soybean plants. Fluorescence-based real-time quantitative PCR (qRT-PCR) showed that the expression level of GmftsH genes varied under different stress treatments. Specifically, the genes within this family exhibited various induction levels in response to stress conditions of 4℃, 20% PEG-6000, and 100 mmol/L NaCl. These findings suggest that the GmftsH gene family may play a crucial role in the abiotic stress response in soybeans. It was also found that the GmftsH7 gene was localized on the cell membrane, and its expression was significantly upregulated under 4 ℃ treatment. In summary, by conducting a genome-wide analysis of the GmftsH gene family, a strong theoretical basis is established for future studies on the functionality of GmftsH genes. CONCLUSIONS: This research can potentially serve as a guide for enhancing the stress tolerance characteristics of soybean.

Enhancement of root sulfur metabolic pathway by overexpression of OAS-TL3 to increase total soybean seed protein content.

Sulfur is essential for plant growth, and the uptake of sulfate by plant roots is the primary source of plant sulfur. Previous studies have shown that the OAS-TL gene is a key enzyme in the sulfur metabolic pathway and regulates cysteine (Cys) synthase production. However, the interaction mechanism of the glycine max OAS-TL3 Cys synthase (OAS-TL3) gene on soybean root morphology construction and seed protein accumulation is unclear. This study shows that mutant M18 has better root growth and development, higher seed protein content, and higher methionine (Met) content in sulfur-containing amino acids than wild-type JN18. By transcriptome sequencing, the differentially expressed OAS-TL3 gene was targeted in the mutant M18 root line. The relative expression of the OAS-TL3 gene in roots, stems, and leaves during the seedling, flowering, and bulking stages of the OAS-TL3 gene overexpression lines is higher than that of the recipient material. Compared to the recipient material JN74, the enzymatic activities, Cys, and GSH contents of OAS-TL are higher in the sulfur metabolic pathway of seedling roots. The receptor material JN74 is exogenously applied with different concentrations of reduced glutathione. The results demonstrate a positive correlation between reduced glutathione on total root length, projected area, surface area, root volume, total root tip number, total bifurcation number, and total crossing number. The Met and total protein contents of sulfur-containing amino acids in soybean seeds of the OAS-TL3 gene overexpression lines are higher than those of the recipient material JN74, while the gene-edited lines show the opposite results. In conclusion, the OAS-TL3 gene positively regulates soybean root growth, root activity, and the content of Met in the seeds through the OAS-TL-Cys-GSH pathway. It breaks the limitation of other amino acids and facilitates the increase of total seed protein content. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-022-01348-y.

The GmXTH1 gene improves drought stress resistance of soybean seedlings.

In order to study the role of GmXTH1 gene in alleviating drought stress, soybean seeds with GmXTH1 gene were transferred by T4 treated with PEG6000 concentration of 0%, 5%, 10%, and 15% respectively. The germination potential, germination rate, germination index, and other indicators were measured. The results showed that the germination potential, germination rate, and germination index of OEA1 and OEA2 strains overexpressed in T4 generation were significantly higher than those of the control material M18. After 0-day, 7-day, and 15-day drought stress, the analysis of seedling phenotypes and root-shoot of different T4 generation transgenic soybean lines showed that under stress conditions, the growth of GmXTH1 overexpression material was generally better than that of the control material M18. The growth of GmXTH1 interference expression material was generally worse than that of the control material M18, with significant differences in plant phenotypes. The root system of GmXTH1 overexpressed material was significantly developed compared with that of the control material M18. The analysis of physiological and biochemical indexes showed that the relative water content and the activity of antioxidant enzymes (superoxide dismutase and peroxidase) of GmXTH1 transgenic soybean material were significantly higher than those of the control material M18, and the accumulation of malondialdehyde was lower under the same stress conditions at seedling stage. Fluorescence quantitative PCR assay showed that the relative expression of GmXTH1 gene in transgenic soybean was significantly increased after drought stress. The results showed that the overexpression of GmXTH1 could increase the total root length, surface area, total projection area, root volume, average diameter, total cross number, and total root tip number, thereby increasing the water intake and reducing the transpiration of water content in leaves, thus reducing the accumulation of MDA and producing more protective enzymes in a more effective and prompt way, reducing cell membrane damage to improve drought resistance of soybean.

Genome-wide association screening and verification of potential genes associated with root architectural traits in maize (Zea mays L.) at multiple seedling stages.

BACKGROUND: Breeding for new maize varieties with propitious root systems has tremendous potential in improving water and nutrients use efficiency and plant adaptation under suboptimal conditions. To date, most of the previously detected root-related trait genes in maize were new without functional verification. In this study, seven seedling root architectural traits were examined at three developmental stages in a recombinant inbred line population (RIL) of 179 RILs and a genome-wide association study (GWAS) panel of 80 elite inbred maize lines through quantitative trait loci (QTL) mapping and genome-wide association study. RESULTS: Using inclusive composite interval mapping, 8 QTLs accounting for 6.44-8.83 % of the phenotypic variation in root traits, were detected on chromosomes 1 (qRDWv3-1-1 and qRDW/SDWv3-1-1), 2 (qRBNv1-2-1), 4 (qSUAv1-4-1, qSUAv2-4-1, and qROVv2-4-1), and 10 (qTRLv1-10-1, qRBNv1-10-1). GWAS analysis involved three models (EMMAX, FarmCPU, and MLM) for a set of 1,490,007 high-quality single nucleotide polymorphisms (SNPs) obtained via whole genome next-generation sequencing (NGS). Overall, 53 significant SNPs with a phenotypic contribution rate ranging from 5.10 to 30.2 % and spread all over the ten maize chromosomes exhibited associations with the seven root traits. 17 SNPs were repeatedly detected from at least two growth stages, with several SNPs associated with multiple traits stably identified at all evaluated stages. Within the average linkage disequilibrium (LD) distance of 5.2 kb for the significant SNPs, 46 candidate genes harboring substantial SNPs were identified. Five potential genes viz. Zm00001d038676, Zm00001d015379, Zm00001d018496, Zm00001d050783, and Zm00001d017751 were verified for expression levels using maize accessions with extreme root branching differences from the GWAS panel and the RIL population. The results showed significantly (P < 0.001) different expression levels between the outer materials in both panels and at all considered growth stages. CONCLUSIONS: This study provides a key reference for uncovering the complex genetic mechanism of root development and genetic enhancement of maize root system architecture, thus supporting the breeding of high-yielding maize varieties with propitious root systems.

Correction to: CRISPR-Cas9 mediated targeted disruption of FAD2-2 microsomal omega-6 desaturase in soybean (Glycine max.L).

An amendment to this paper has been published and can be accessed via the original article.

Genome-wide association study identifies candidate genes related to oleic acid content in soybean seeds.

BACKGROUND: Soybean oil is a complex mixture of five fatty acids (palmitic, stearic, oleic, linoleic, and linolenic). Soybean oil with a high oleic acid content is desirable because this monounsaturated fatty acid improves the oxidative stability of the oil. To investigate the genetic architecture of oleic acid in soybean seeds, 260 soybean germplasms from Northeast China were collected as natural populations. A genome-wide association study (GWAS) was conducted on a panel of 260 germplasm resources. RESULTS: Phenotypic identification results showed that the oleic acid content varied from 8.2 to 35.0%. A total of 2,311,337 single-nucleotide polymorphism (SNP) markers were obtained. GWAS analysis showed that there were many genes related to oleic acid content with a contribution rate of 7%. The candidate genes Glyma.11G229600.1 on chromosome 11 and Glyma.04G102900.1 on chromosome 4 were detected in a 2-year-long GWAS. The candidate gene Glyma.11G229600.1 showed a positive correlation with the oleic acid content, and the correlation coefficient was 0.980, while Glyma.04G102900.1 showed a negative correlation, with a coefficient of - 0.964. CONCLUSIONS: Glyma.04G102900.1 on chromosome 4 and Glyma.11G229600.1 on chromosome 11 were detected in both analyses (2018 and 2019). Glyma.04G102900.1 and Glyma.11G229600.1 are new key candidate genes related to oleic acid in soybean seeds. These results will be useful for high-oleic soybean breeding.

Construction and Analysis of GmFAD2-1A and GmFAD2-2A Soybean Fatty Acid Desaturase Mutants Based on CRISPR/Cas9 Technology.

The soybean fatty acid desaturase family is composed of seven genes, but the function of each gene has not been reported. Bioinformatics was used to analyse the structure of genes in this family, as well as the correlation between Δ12-fatty acid desaturase II (FAD2) expression and oleic acid content on different days after flowering of soybean. In the present study, CRISPR/Cas9 technology was used to construct single and double mutant knockout vectors of functional genes in the FAD2 family. Analysis of the molecular biology and expression patterns of genes in the FAD2 family, namely, GmFAD2-1A (Glyma.10G278000) and GmFAD2-2A (Glyma.19G147300), showed that they had little homology with other soybean FAD2 genes, and that their function was slightly changed. Sequencing of the target showed that the editing efficiency of the GmFAD2-1A and GmFAD2-2A genes was 95% and 55.56%, respectively, and that the double mutant editing efficiency was 66.67%. The mutations were divided into two main types, as follows: base deletion and insertion. A near-infrared grain analyser determined the following results: In the T2 generation, the oleic acid content increased from 17.10% to 73.50%; the linoleic acid content decreased from 62.91% to 12.23%; the protein content increased from 37.69% to 41.16%; in the T3 generation, the oleic acid content increased from 19.15% to 72.02%; the linoleic acid content decreased from 56.58% to 17.27%. In addition, the protein content increased from 37.52% to 40.58% compared to that of the JN38 control variety.

Soybean plants expressing the Bacillus thuringiensis cry8-like gene show resistance to Holotrichia parallela.

BACKGROUND: Cry8-like from Bacillus thuringiensis (Bt) encodes an insecticidal crystal (Cry) protein. Holotrichia parallela (Coleoptera: Scarabaeoidae), commonly known as the dark black chafer, is a troublesome pest of soybean (Glycine max). To test whether cry8-like can confer resistance against H. parallela to soybean, we introduced cry8-like from the Bt strain HBF-18 into soybean cultivar Jinong 28. RESULTS: Quantitative reverse transcription-PCR analysis demonstrated that cry8-like was expressed most highly in soybean leaves. In addition, Southern blot assays revealed that one copy of the integrated fragment was present in the transformed plants. Eight independent cry8-like transgenic lines were subsequently fed on by H. parallela. Under H. parallela feeding stress, the survival rates of the non-transgenic plants were 92% lower than those of the transgenic plants. The mortality rate of H. parallela increased when the larvae fed on the roots of T1 transgenic soybean plants. Moreover, the surviving larvae were deformed, and their growth was inhibited. CONCLUSIONS: Collectively, our data suggest that transgenic soybean plants expressing the cry8-like gene are more resistant to H. parallela than non-transgenic plants and that transgenic expression of the cry8-like gene may represent a promising strategy for engineering pest tolerance. The events generated in this study could thus be utilized in soybean breeding programs.

CRISPR-Cas9 mediated targeted disruption of FAD2-2 microsomal omega-6 desaturase in soybean (Glycine max.L).

BACKGROUND: Recent innovation in the field of genome engineering encompasses numerous levels of plant genome engineering which attract the substantial excitement of plant biologist worldwide. RNA-guided CRISPR Cas9 system has appeared a promising tool in site-directed mutagenesis due to its innovative utilization in different branches of biology. CRISPR-Cas9 nuclease system have supersedes all previously existed strategies and their associated pitfalls encountered with site-specific mutagenesis. RESULTS: Here we demonstrated an efficient sequence specific integration/mutation of FAD2-2 gene in soybean using CRISPR-Cas9 nuclease system. A single guided RNA sequence was designed with the help of a number of bioinformatics tools aimed to target distinct sites of FAD2-2 loci in soybean. The binary vector (pCas9-AtU6-sgRNA) has been successfully transformed into soybean cotyledon using Agrobacterium tumafacien. Taken together our findings complies soybean transgenic mutants subjected to targeted mutation were surprisingly detected in our target gene. Furthermore, the detection of Cas9 gene, BAR gene, and NOS terminator were carried out respectively. Southern blot analysis confirmed the stable transformation of Cas9 gene into soybean. Real time expression with qRT-PCR and Sanger sequencing analysis confirmed the efficient CRISPR-Cas9/sgRNA induced mutation within the target sequence of FAD2-2 loci. The integration of FAD2-2 target region in the form of substitution, deletions and insertions were achieved with notably high frequency and rare off-target mutagenesis. CONCLUSION: High frequent mutation efficiency was recorded as 21% out of all transgenic soybean plants subjected to targeted mutagenesis. Furthermore, Near-infrared spectroscopy (NIR) indicates the entire fatty acid profiling obtained from the mutants seeds of soybean. A considerable modulation in oleic acid content up to (65.58%) whereas the least level of linoleic acid is (16.08%) were recorded. Based on these finding CRISPR-Cas9 system can possibly sum up recent development and future challenges in producing agronomically important crops.

Function of chalcone reductase gene CHR1 in soybean.

To verify the function of chalcone reductase gene (CHR1) in soybean Daidzein synthesis, CHR1 gene in soybean was cloned, and an RNAi expression vector pCPB-CHR1-RNAi was constructed. Four transformed plants in T0 generation and thirteen transformed plants in T1 generation of soybean "Jinong28" were obtained by agrobacterium-mediated genetic transformation, in which transcriptions of CHR1 gene were depressed. Southern blotting showed the functional fragment of pCPB-CHR1-RNAi was integrated into the genome of recipient soybean in the form of a single copy. Detection of the transcription of CHR1 gene using quantitative real-time PCR (qRT-PCR) showed that the expression of CHR1 gene in transformed plants decreased 60%-99% compared to the recipient soybean, while the content of isoliquiritigenin, the precursors of daidzein, decreased 38.7%. These results indicate that RNA interference can suppress the transcription of CHR1 gene expression successfully.

Screening and Validation of Leaf Width-Related Genes in Inbred Maize Lines.

Leaf width is a key determinant of planting density and photosynthetic efficiency. In an effort to determine which genes regulate maize plant leaf width, we performed a genome-wide association study (GWAS) of 1.49 × 106 single nucleotide polymorphisms (SNPs) in 80 sequenced backbone inbred maize lines in Jilin Province, China, based upon phenotypic leaf width data from two years. In total, 14 SNPs were identified as being significantly related to leaf width (p < 0.000001), with these SNPs being located on chromosomes 1, 2, 3, 5, 6, 7, 8, and 9. A total of five candidate genes were identified within a mean linkage disequilibrium (LD) distance of 9.7 kb, with a significant SNP being identified within the Zm00001d044327 candidate gene. RNA was then isolated from 12 different inbred maize lines from this GWAS study cohort and was used to conduct qPCR analyses which revealed significant differences in Zm00001d044327 expression among strains exhibiting significant differences in leaf width. Based on an assessment of EMS mutant lines harboring a conserved amino acid stop mutation and two non-synonymous mutations in Zm00001d044327 that exhibited a narrow leaf width, these data suggested that Zm00001d044327 is a key regulator of maize leaf width.

Cloning of the Soybean GmNHL1 Gene and Functional Analysis under Salt Stress.

When encountered in the soybean seedling stage, salt stress has serious impacts on plant growth and development. This study explores the role of the soybean NDR1/HIN1-like family gene GmNHL1 under salt stress. First, the GmNHL1 gene was successfully cloned, and bioinformatic analysis revealed multiple cis-acting elements which are related to adversity stress and involved in the oxidative response in the promoter region. Sub-cellular localization analysis indicated that the protein expressed by GmNHL1 was localized on the cell membrane. An over-expression vector of the target gene and a CRISPR/Cas9 gene-editing vector were constructed, and the recipient soybean variety Jinong 74 was genetically transformed using the Agrobacterium tumefaciens-mediated method. By analyzing the performance of the different plants under salt stress, the results showed that GmNHL1 was over-expressed in the T2 generation. The germination potential, germination rate, germination index, and vitality index of the strain were significantly higher than those of the recipient control JN74. Under salt stress conditions, the root microanatomical structure of the GmNHL1 over-expressing material remained relatively intact, and its growth was better than that of the recipient control JN74. Measurement of physiological and biochemical indicators demonstrated that, compared with the receptor control JN74, the malondialdehyde and O2- contents of the GmNHL1 over-expressing material were significantly reduced, while the antioxidant enzyme activity, proline content, and chlorophyll content significantly increased; however, the results for GmNHL1 gene-edited materials were the opposite. In summary, over-expression of GmNHL1 can improve the salt tolerance of plants and maintain the integrity of the root anatomical structure, thereby more effectively and rapidly reducing the accumulation of malondialdehyde and O2- content and increasing antioxidant enzyme activity. This reduces cell membrane damage, thereby improving the salt tolerance of soybean plants. These results help to better understand the mechanism of salt tolerance in soybean plants, laying a theoretical foundation for breeding new stress-resistant varieties of soybean.

Identification of the Function of the Pathogenesis-Related Protein GmPR1L in the Resistance of Soybean to Cercospora sojina Hara.

Pathogenesis-related proteins, often used as molecular markers of disease resistance in plants, can enable plants to obtain systemic resistance. In this study, a gene encoding a pathogenesis-related protein was identified via RNA-seq sequencing analysis performed at different stages of soybean seedling development. Because the gene sequence showed the highest similarity with PR1L sequence in soybean, the gene was named GmPR1-9-like (GmPR1L). GmPR1L was either overexpressed or silenced in soybean seedlings through Agrobacterium-mediated transformation to examine the resistance of soybean to infection caused by Cercospora sojina Hara. The results revealed that GmPR1L-overexpressing soybean plants had a smaller lesion area and improved resistance to C. sojina infection, whereas GmPR1L-silenced plants had low resistance to C. sojina infection. Fluorescent real-time PCR indicated that overexpression of GmPR1L induced the expression of genes such as WRKY, PR9, and PR14, which are more likely to be co-expressed during C. sojina infection. Furthermore, the activities of SOD, POD, CAT, and PAL were significantly increased in GmPR1L-overexpressing soybean plants after seven days of infection. The resistance of the GmPR1L-overexpressing lines OEA1 and OEA2 to C. sojina infection was significantly increased from a neutral level in wild-type plants to a moderate level. These findings predominantly reveal the positive role of GmPR1L in inducing resistance to C. sojina infection in soybean, which may facilitate the production of improved disease-resistant soybean cultivars in the future.

Overexpression of TCP9-like gene enhances salt tolerance in transgenic soybean.

TEOSINTE BRANCHED1/CYCLOIDEA/PROLIFERATING CELL FACTOR (TCP) transcription factors are a plant-specific family and play roles in plant growth, development, and responses to biotic and abiotic stresses. However, little is known about the functions of the TCP transcription factors in the soybean cultivars with tolerance to salt stress. In this study, TCP9-like, a TCP transcription factor, was identified in the soybean cultivars exposed to salt stress. The expression of TCP9-like gene in the roots of salt-tolerant soybean cultivars was higher than that in salt-sensitive cultivars treated with NaCl. The overexpression of TCP9-like enhanced the salt tolerance of the salt-sensitive soybean cultivar 'DN50'. In T2 generation, the plants with TCP9-like overexpression had significantly lower Na+ accumulation and higher K+ accumulation than the WT plants exposed to 200 or 250 mmol/L NaCl. The K+/Na+ ratio in the plants overexpressing TCP9-like was significantly higher than that in WT plants treated with 200 mmol/L NaCl. Meanwhile, the overexpression of TCP9-like up-regulated the expression levels of GmNHX1, GmNHX3, GmSOS1, GmSOS2-like, and GmHKT1, which were involved in the K+/Na+ homeostasis pathway. The findings indicated that TCP9-like mediated the regulation of both Na+ and K+ accumulation to improve the tolerance of soybean to salt stress.

[Cloning and functional analysis of SCTF-1 encoding a C2H2-type Zinc finger protein from soybean].

The zinc finger protein is one of the proteins with finger-like domain. Some of them are transcription factors which play important role in plant growth and plant resistance to abiotic stresses. In this paper, a novel C2H2-type zinc finger protein gene SCTF-1 (GenBank accession number JQ692081) was isolated from soybean (Glycine max (L.) Merr.) This gene has a 699 bp ORF (open reading frame) with no intron and encodes a 24.9 kDa protein with 233 amino acids. Its isoelectric point (pI) is 8.33. The SCTF-1 protein contains two typical C2H2-type zinc finger domains. Both of them have highly conserved amino acid sequence-QALGGH which is a particular characteristic of plant. Transient expression of the GFP-SCTF-1 protein in onion epidermal cell showed that SCTF-1 was localized in cell nuclei. RT-PCR results showed that SCTF-1 gene was expressed with high levels in flowers and leaves in soybean, but low in roots and stems. The expression of SCTF-1 gene was strongly induced by low temperature in the soybean seedlings. Overexpression of SCTF-1 enhanced cold tolerance of transgenic tobacco (Nicotiana tabacum L.) compared to the control.

CRISPR/Cas9-mediated targeted mutagenesis of GmTCP19L increasing susceptibility to Phytophthora sojae in soybean.

The TEOSINTE BRANCHED1/CYCLOIDEA/PROLIFERATING CELL FACTOR (TCP) transcription factors is one of the superfamilies of plant-specific transcription factors involved in plant growth, development, and biotic and abiotic stress. However, there is no report on the research of the TCP transcription factors in soybean response to Phytophthora sojae. In this study, Agrobacterium-mediated transformation was used to introduce the CRISPR/Cas9 expression vector into soybean cultivar "Williams 82" and generated targeted mutants of GmTCP19L gene, which was previously related to involve in soybean responses to P. sojae. We obtained the tcp19l mutants with 2-bp deletion at GmTCP19L coding region, and the frameshift mutations produced premature translation termination codons and truncated GmTCP19L proteins, increasing susceptibility to P. sojae in the T2-generation. These results suggest that GmTCP19L encodes a TCP transcription factor that affects plant defense in soybean. The new soybean germplasm with homozygous tcp19l mutations but the BAR and Cas9 sequences were undetectable using strip and PCR methods, respectively, suggesting directions for the breeding or genetic engineering of disease-resistant soybean plants.

Fine Mapping and Functional Research of Key Genes for Photoperiod Sensitivity in Maize.

Maize is native to the tropics and is very sensitive to photoperiod. Planting in temperate regions with increased hours of daylight always leads to late flowering, sterility, leggy plants, and increased numbers of maize leaves. This phenomenon severely affects the utilization of tropical maize germplasm resources. The sensitivity to photoperiod is mainly reflected in differences in plant height (PH), ear height (EH), total leaf number (LN), leaf number under ear (LE), silking stage (SS), and anthesis stage (AT) in the same variety under different photoperiod conditions. These differences are more pronounced for varieties that are more sensitive to photoperiod. In the current study, a high-density genetic map was constructed from a recombinant inbred line (RIL) population containing 209 lines to map the quantitative trait loci (QTL) for photoperiod sensitivity of PH, EH, LN, LE, SS, and AT. A total of 39 QTL were identified, including three consistent major QTL. We identified candidate genes in the consensus major QTL region by combined analysis of transcriptome data, and after enrichment by GO and KEGG, we identified a total of four genes (Zm00001d006212, Zm00001d017241, Zm00001d047761, and Zm00001d047632) enriched in the plant circadian rhythm pathway (KEGG:04712). We analyzed the expression levels of these four genes, and the analysis results showed that there were significant differences in response under different photoperiod conditions for three of them (Zm00001d047761, Zm00001d006212 and Zm00001d017241). The results of functional verification showed that the expression patterns of genes rhythmically oscillated, which can affect the length of the hypocotyl and the development of the shoot apical meristem. We also found that the phenotypes of the positive plants were significantly different from the control plants when they overexpressed the objective gene or when it was knocked out, and the expression period, phase, and amplitude of the target gene also shifted. The objective gene changed its own rhythmic oscillation period, phase, and amplitude with the change in the photoperiod, thereby regulating the photoperiod sensitivity of maize. These results deepen our understanding of the genetic structure of photoperiod sensitivity and lay a foundation for further exploration of the regulatory mechanism of photoperiod sensitivity.

Genome-Wide Association Study of Root and Shoot Related Traits in Spring Soybean (Glycine max L.) at Seedling Stages Using SLAF-Seq.

Root systems can display variable genetic architectures leading to nutrient foraging or improving abiotic stress tolerance. Breeding for new soybean varieties with efficient root systems has tremendous potential in enhancing resource use efficiency and plant adaptation for challenging climates. In this study, root related traits were analyzed in a panel of 260 spring soybean with genome-wide association study (GWAS). Genotyping was done with specific locus amplified fragment sequencing (SLAF-seq), and five GWAS models (GLM, MLM, CMLM, FaST-LMM, and EMMAX) were used for analysis. A total of 179,960 highly consistent SNP markers distributed over the entire genome with an inter-marker distance of 2.36 kb was used for GWAS analysis. Overall, 27 significant SNPs with a phenotypic contribution ranging from 20 to 72% and distributed on chromosomes 2, 6, 8, 9, 13, 16 and 18 were identified and two of them were found to be associated with multiple root-related traits. Based on the linkage disequilibrium (LD) distance of 9.5 kb for the different chromosomes, 11 root and shoot regulating genes were detected based on LD region of a maximum 55-bp and phenotypic contribution greater than 22%. Expression analysis revealed an association between expression levels of those genes and the degree of root branching number. The current study provides new insights into the genetic architecture of soybean roots, and the underlying SNPs/genes could be critical for future breeding of high-efficient root system in soybean.

Identification and Functional Verification of Cold Tolerance Genes in Spring Maize Seedlings Based on a Genome-Wide Association Study and Quantitative Trait Locus Mapping.

Maize (Zea mays L.) is a tropical crop, and low temperature has become one of the main abiotic stresses for maize growth and development, affecting many maize growth processes. The main area of maize production in China, Jilin province, often suffers from varying degrees of cold damage in spring, which seriously affects the quality and yield of maize. In the face of global climate change and food security concerns, discovering cold tolerance genes, developing cold tolerance molecular markers, and creating cold-tolerant germplasm have become urgent for improving maize resilience against these conditions and obtaining an increase in overall yield. In this study, whole-genome sequencing and genotyping by sequencing were used to perform genome-wide association analysis (GWAS) and quantitative trait locus (QTL) mapping of the two populations, respectively. Overall, four single-nucleotide polymorphisms (SNPs) and 12 QTLs were found to be significantly associated with cold tolerance. Through joint analysis, an intersection of GWAS and QTL mapping was found on chromosome 3, on which the Zm00001d002729 gene was identified as a potential factor in cold tolerance. We verified the function of this target gene through overexpression, suppression of expression, and genetic transformation into maize. We found that Zm00001d002729 overexpression resulted in better cold tolerance in this crop. The identification of genes associated with cold tolerance contributes to the clarification of the underlying mechanism of this trait in maize and provides a foundation for the adaptation of maize to colder environments in the future, to ensure food security.

Genetic mapping of northern corn leaf blight-resistant quantitative trait loci in maize.

Northern corn leaf blight (NCLB), a corn disease infected by Exserohilum turcicum, can cause loss of harvest and economy. Identification or evaluation of NCLB-resistant quantitative trait loci (QTL) and genes could improve maize breeds. This study aimed to identify novel QTLs for NCLB-resistance.Two maize strains (BB and BC) were utilized to generate B73 × B97 and B73 × CML322 and constructed the genetic linkage using high-throughput single nucleotide polymorphism (SNP) linkage map analysis of 170 (BB) and 163(BC) recombinant inbred line (RIL) genomic DNA samples. NCLB-resistant QTL was associated with phenotypic data from the field trial of 170 BB and 163 BC strains over two years using these 1100 SNPs to identify high-density NCLB-resistant QTLs.In BB, QTL of the NCLB resistance was on chromosome 1 and 3 (LOD scores between 2.74 and 5.44); in BC, QTL of NCLB resistance was on chromosome 1, 2, 4, 8, and 9 (LOD scores between 2.52 and 8.53). A number of genes or genetic information related to NCLB resistance in both BB and BC were identified with the maximum number of genes/NCLB resistance-related QTL on chromosome 3 for BB and on chromosome 1 for BC.This study successfully mapped and identified NCLB-resistant QTL and genes for these 2 different maize strains, which provides insightful information for future study of NCLB-resistance and selection of NCLB-resistant maize variants.