Illegitimate Recombination between Duplicated Genes Generated from Recursive Polyploidizations Accelerated the Divergence of the Genus Arachis

The peanut (Arachis hypogaea L.) is the leading oil and food crop among the legume family. Extensive duplicate gene pairs generated from recursive polyploidizations with high sequence similarity could result from gene conversion, caused by illegitimate DNA recombination. Here, through synteny-based comparisons of two diploid and three tetraploid peanut genomes, we identified the duplicated genes generated from legume common tetraploidy (LCT) and peanut recent allo-tetraploidy (PRT) within genomes. In each peanut genome (or subgenomes), we inferred that 6.8–13.1% of LCT-related and 11.3–16.5% of PRT-related duplicates were affected by gene conversion, in which the LCT-related duplicates were the most affected by partial gene conversion, whereas the PRT-related duplicates were the most affected by whole gene conversion. Notably, we observed the conversion between duplicates as the long-lasting contribution of polyploidizations accelerated the divergence of different Arachis genomes. Moreover, we found that the converted duplicates are unevenly distributed across the chromosomes and are more often near the ends of the chromosomes in each genome. We also confirmed that well-preserved homoeologous chromosome regions may facilitate duplicates’ conversion. In addition, we found that these biological functions contain a higher number of preferentially converted genes, such as catalytic activity-related genes. We identified specific domains that are involved in converted genes, implying that conversions are associated with important traits of peanut growth and development.

Genome-wide identification and evolutionary analysis of RLKs involved in the response to aluminium stress in peanut

Background As an important cash crop, the yield of peanut is influenced by soil acidification and pathogen infection. Receptor-like protein kinases play important roles in plant growth, development and stress responses. However, little is known about the number, location, structure, molecular phylogeny, and expression of RLKs in peanut, and no comprehensive analysis of RLKs in the Al stress response in peanuts have been reported. Results A total of 1311 AhRLKs were identified from the peanut genome. The AhLRR-RLKs and AhLecRLKs were further divided into 24 and 35 subfamilies, respectively. The AhRLKs were randomly distributed across all 20 chromosomes in the peanut. Among these AhRLKs, 9.53% and 61.78% originated from tandem duplications and segmental duplications, respectively. The ka/ks ratios of 96.97% (96/99) of tandem duplication gene pairs and 98.78% (646/654) of segmental duplication gene pairs were less than 1. Among the tested tandem duplication clusters, there were 28 gene conversion events. Moreover, all total of 90 Al-responsive AhRLKs were identified by mining transcriptome data, and they were divided into 7 groups. Most of the Al-responsive AhRLKs that clustered together had similar motifs and evolutionarily conserved structures. The gene expression patterns of these genes in different tissues were further analysed, and tissue-specifically expressed genes, including 14 root-specific Al-responsive AhRLKs were found. In addition, all 90 Al-responsive AhRLKs which were distributed unevenly in the subfamilies of AhRLKs, showed different expression patterns between the two peanut varieties (Al-sensitive and Al-tolerant) under Al stress. Conclusions In this study, we analysed the RLK gene family in the peanut genome. Segmental duplication events were the main driving force for AhRLK evolution, and most AhRLKs subject to purifying selection. A total of 90 genes were identified as Al-responsive AhRLKs, and the classification, conserved motifs, structures, tissue expression patterns and predicted functions of Al-responsive AhRLKs were further analysed and discussed, revealing their putative roles. This study provides a better understanding of the structures and functions of AhRLKs and Al-responsive AhRLKs.

Identification of consistent QTL for time to maturation in Virginia-type Peanut (Arachis hypogaea L.)

Background Time-to-maturation (TTM) is an important trait contributing to adaptability, yield and quality in peanut (Arachis hypogaea L). Virginia market-type peanut belongs to the late-maturing A. hypogaea subspecies with considerable variation in TTM within this market type. Consequently, planting and harvesting schedule of peanut cultivars, including Virginia market-type, need to be optimized to maximize yield and grade. Little is known regarding the genetic control of TTM in peanut due to the challenge of phenotyping and limited DNA polymorphism. Here, we investigated the genetic control of TTM within the Virginia market-type peanut using a SNP-based high-density genetic map. A recombinant inbred line (RIL) population, derived from a cross between two Virginia-type cultivars ‘Hanoch’ and ‘Harari’ with contrasting TTM (12–15 days on multi-years observations), was phenotyped in the field for 2 years following a randomized complete block design. TTM was estimated by maturity index (MI). Other agronomic traits like harvest index (HI), branching habit (BH) and shelling percentage (SP) were recorded as well. Results MI was highly segregated in the population, with 13.3–70.9% and 28.4–80.2% in years 2018 and 2019. The constructed genetic map included 1833 SNP markers distributed on 24 linkage groups, covering a total map distance of 1773.5 cM corresponding to 20 chromosomes on the tetraploid peanut genome with 1.6 cM mean distance between the adjacent markers. Thirty QTL were identified for all measured traits. Among the four QTL regions for MI, two consistent QTL regions (qMIA04a,b and qMIB03a,b) were identified on chromosomes A04 (118680323–125,599,371; 6.9Mbp) and B03 (2839591–4,674,238; 1.8Mbp), with LOD values of 5.33–6.45 and 5–5.35 which explained phenotypic variation of 9.9–11.9% and 9.3–9.9%, respectively. QTL for HI were found to share the same loci as MI on chromosomes B03, B05, and B06, demonstrating the possible pleiotropic effect of HI on TTM. Significant but smaller effects on MI were detected for BH, pod yield and SP. Conclusions This study identified consistent QTL regions conditioning TTM for Virginia market-type peanut. The information and materials generated here can be used to further develop molecular markers to select peanut idiotypes suitable for diverse growth environments.

Genome‐wide identification of mlo genes in the cultivated peanut (Arachis hypogaea L.)

Powdery mildew disease caused by Oidium arachidis poses a threat to peanut production in Africa. Loss of function mutants of specific Mlo (Mildew Locus O) genes have provided broad-spectrum and durable resistance against pathogen in many crop species. Since there is huge potential to utilize susceptibility gene-mediated resistance in crop improvement, genome-wide mining of susceptibility genes is required for further researches. However, the susceptibility genes have not been characterized in peanut genome. In this research study, the genome of the cultivated peanut was used as reference to identify the AhMlo loci. Our results revealed that 25 AhMlo loci were identified and distributed on the chromosomes of the cultivated peanut. Eleven AhMlo loci were located on the A-genome while the remaining 14 on the B-genome. Variable number of inserted intron sequences (4–14) and transmembrane helix (4–8) were observed in the coding sequence of the AhMlo loci. Furthermore, phylogenetic analysis of the AhMlo loci along with homologs from other species has clustered the AhMlo loci into six clades. Three AhMlo loci were clustered in the clade V known to regroup the powdery susceptibility loci in dicots. Additionally, four core promoters were predicted on the promoter region of the specific AhMlo along with cis-regulatory elements related to PM susceptibility. These results provided strong evidence of the identification and distribution of the Mlo loci in the cultivated peanut genome and the identified specific AhMlo loci can be used for loss of susceptibility study.

Genome-wide identification and evolutionary analysis of RLKs involved in response to Aluminum stress in peanut

Background: As an important cash crop, the yield of peanut is influenced by soil acidification and pathogen infection. Receptor-like protein kinase plays important roles in plant growth, development and stress responses. However, little is known about the number, location, structure, molecular phylogenetic, and expression of the RLKs in peanut, and no comprehensive analysis of RLKs in Al stress response in peanut have been reported. Results: A total of 1311 AhRLKs were identified from the peanut genome. The AhLRR-RLKs and AhLec-RLKs were further divided into 24 subfamilies and 35 subfamilies, respectively. The AhRLKs are randomly distributed across all 20 chromosomes in peanut. Among them, 67.8% and 0.6% of the AhRLKs originated from tandem duplications and segmental duplications, respectively. The ka/ks ratios of 94.9% (1290/1360) of AhRLKs were less than 1. Moreover, totally 90 Al-responsive AhRLKs were identified by mining transcriptome data, and they were divided into 7 groups. Most of Al responsive AhRLKs clustered together had similar motifs and evolutionarily conserved structures. The gene expression patterns of these genes in different tissues were further analyzed, and tissue specific expression genes, including 14 root-specific Al responsive AhRLKs were found. Besides, all of the 90 Al responsive AhRLKs which distributed unevenly in the subfamilies of AhRLKs have different expression pattern between two peanut varieties (Al-sensitive and Al-tolerant) under Al stress.Conclusions: In this study, we analyzed the RLK gene family by the peanut genome. Tandem replication events were the main driving force for AhRLKs evolution, and most AhRLKs were selected for purification. A total of 90 genes were identified as Al responsive AhRLKs, and the classification, conservative motif, structure, tissue expression pattern and predicted function of Al responsive AhRLKs were further analyzed and discussed, revealing their putative roles. This study provides a better understanding of the structures and functions of AhRLKs as well as Al responsive AhRLKs.

Pod and Seed Trait QTL Identification To Assist Breeding for Peanut Market Preferences

Although seed and pod traits are important for peanut breeding, little is known about the inheritance of these traits. A recombinant inbred line (RIL) population of 156 lines from a cross of Tifrunner x NC 3033 was genotyped with the Axiom_Arachis1 SNP array and SSRs to generate a genetic map composed of 1524 markers in 29 linkage groups (LG). The genetic positions of markers were compared with their physical positions on the peanut genome to confirm the validity of the linkage map and explore the distribution of recombination and potential chromosomal rearrangements. This linkage map was then used to identify Quantitative Trait Loci (QTL) for seed and pod traits that were phenotyped over three consecutive years for the purpose of developing trait-associated markers for breeding. Forty-nine QTL were identified in 14 LG for seed size index, kernel percentage, seed weight, pod weight, single-kernel, double-kernel, pod area and pod density. Twenty QTL demonstrated phenotypic variance explained (PVE) greater than 10% and eight more than 20%. Of note, seven of the eight major QTL for pod area, pod weight and seed weight (PVE >20% variance) were attributed to NC 3033 and located in a single linkage group, LG B06_1. In contrast, the most consistent QTL for kernel percentage were located on A07/B07 and derived from Tifrunner.

The Family of Peanut Fatty Acid Desaturase Genes and a Functional Analysis of Four ω-3 AhFAD3 Members

The synthesis of α-linolenic acid (ALA) requires the activity of ω-3 fatty acid desaturases (ω-3 FADs). The quality of peanut oil would be much improved if the content of ALA could be increased. A scan of the peanut genome revealed that it harbored 36 FAD genes, mapping to 16 of the species’ 20 chromosomes. A phylogenetic analysis concluded that these genes belonged to six sub-families, namely stearoyl-acyl-acyl carrier protein desaturases (SAD), FAD2, FAD3, FAD4/5, FAD6 and FAD7/8. Of these, FAD3 and FAD7/8 encoded ω-3 FADs, while genes belonging to the other four sub-families encoded ω-6 FADs. Based on RNA-Seq data, each of the 36 FAD genes was shown to be transcribed in non-stressed plants, but there was variation between them with respect to which organs they were transcribed in. Four ω-3 AhFAD3 genes were functionally characterized; when expressed in Arabidopsis thaliana protoplasts, each was localized mainly in the endoplasmic reticulum, while within peanut, the genes were more strongly transcribed in the developing seed than in either the root or the leaf. When constitutively expressed in Arabidopsis thaliana, both the total fatty acid content of the seed and the relative contribution of ALA were increased. The transgenic seedlings also exhibited an improved level of survival when challenged by salinity stress.

QTL identification for seed weight and size based on a high-density SLAF-seq genetic map in peanut (Arachis hypogaea L.)

Background The cultivated peanut is an important oil and cash crop grown worldwide. To meet the growing demand for peanut production each year, genetic studies and enhanced selection efficiency are essential, including linkage mapping, genome-wide association study, bulked-segregant analysis and marker-assisted selection. Specific locus amplified fragment sequencing (SLAF-seq) is a powerful tool for high density genetic map (HDGM) construction and quantitative trait loci (QTLs) mapping. In this study, a HDGM was constructed using SLAF-seq leading to identification of QTL for seed weight and size in peanut. Results A recombinant inbred line (RIL) population was advanced from a cross between a cultivar ‘Huayu36’ and a germplasm line ‘6–13’ with contrasting seed weight, size and shape. Based on the cultivated peanut genome, a HDGM was constructed with 3866 loci consisting of SLAF-seq and simple sequence repeat (SSR) markers distributed on 20 linkage groups (LGs) covering a total map distance of 1266.87 cM. Phenotypic data of four seed related traits were obtained in four environments, which mostly displayed normal distribution with varied levels of correlation. A total of 27 QTLs for 100 seed weight (100SW), seed length (SL), seed width (SW) and length to width ratio (L/W) were identified on 8 chromosomes, with LOD values of 3.16–31.55 and explaining phenotypic variance (PVE) from 0.74 to 83.23%. Two stable QTL regions were identified on chromosomes 2 and 16, and gene content within these regions provided valuable information for further functional analysis of yield component traits. Conclusions This study represents a new HDGM based on the cultivated peanut genome using SLAF-seq and SSRs. QTL mapping of four seed related traits revealed two stable QTL regions on chromosomes 2 and 16, which not only facilitate fine mapping and cloning these genes, but also provide opportunity for molecular breeding of new peanut cultivars with improved seed weight and size.

Pod and seed trait QTL identification to assist breeding for peanut market preferences

ABSTRACTAlthough seed and pod traits are important for peanut breeding, little is known about the inheritance of these traits. A recombinant inbred line (RIL) population of 156 lines from a cross of Tifrunner x NC 3033 was genotyped with the Axiom_Arachis1 SNP array and SSRs to generate a genetic map composed of 1524 markers in 29 linkage groups (LG). The genetic positions of markers were compared with their physical positions on the peanut genome to confirm the validity of the linkage map and explorethe distribution of recombination and potential chromosomal rearrangements. These traits were phenotyped over three consecutive years for the purpose of developing trait-associated markers for breeding. Forty-nine QTL were identified in 14 LG for seed size index, kernel percentage, seed weight, pod weight, single-kernel, double-kernel, pod area and pod density. Twenty QTL demonstrated phenotypic variance explained (PVE) greater than 10% and eight more than 20%. Of note, seven of the eight major QTL for pod area, pod weight and seed weight (PVE >20% variance) were attributed to NC 3033 and located in a single linkage group, LG B06_1. In contrast, the most consistent QTL for kernel percentage were located on A07/B07 and derived from Tifrunner.