Boshou Liao

The genome sequences of Arachis duranensis and Arachis ipaensis , the diploid ancestors of cultivated peanut

Cultivated peanut (Arachis hypogaea) is an allotetraploid with closely related subgenomes of a total size of ∼2.7 Gb. This makes the assembly of chromosomal pseudomolecules very challenging. As a foundation to understanding the genome of cultivated peanut, we report the genome sequences of its diploid ancestors (Arachis duranensis and Arachis ipaensis). We show that these genomes are similar to cultivated peanuts A and B subgenomes and use them to identify candidate disease resistance genes, to guide tetraploid transcript assemblies and to detect genetic exchange between cultivated peanuts subgenomes. On the basis of remarkably high DNA identity of the A. ipaensis genome and the B subgenome of cultivated peanut and biogeographic evidence, we conclude that A. ipaensis may be a direct descendant of the same population that contributed the B subgenome to cultivated peanut.

Comparative Assessment of Genetic Diversity of Peanut (Arachis hypogaea L.) Genotypes with Various Levels of Resistance to Bacterial Wilt Through SSR and AFLP Analyses

Bacterial wilt (BW) caused by Ralstonia solanacearum is an important constraint to peanut (Arachis hypogaea L.) production in several Asian and African countries, and planting BW-resistant cultivars is the most feasible method for controlling the disease. Although several BW-resistant peanut germplasm accessions have been identified, the genetic diversity among these has not been properly investigated, which has impeded efficient utilization. In this study, the genetic relationships of 31 peanut genotypes with various levels of resistance to BW were assessed based on SSR and AFLP analyses. Twenty-nine of 78 SSR primers and 32 of 126 AFLP primer combinations employed in this study were polymorphic amongst the peanut genotypes tested. The SSR primers amplified 91 polymorphic loci in total with an average of 3.14 alleles per primer, and the AFLP primers amplified 72 polymorphic loci in total with an average of 2.25 alleles per primer. Four SSR primers (14H06, 7G02, 3A8, 16C6) and one AFLP primer (P1M62) were found to be most efficient in detecting diversity. The genetic distance between pairs of genotypes ranged from 0.12 to 0.94 with an average of 0.53 in the SSR data and from 0.06 to 0.57 with an average of 0.25 in the AFLP data. The SSR-based estimates of the genetic distance were generally larger than that based on the AFLP data. The genotypes belonging to subsp. fastigiata possessed wider diversity than that of subsp. hypogaea. The clustering of genotypes based on the SSR and AFLP data were similar but the SSR clustering was more consistent with morphological classification of A. hypogaea. Optimum diverse genotypes of both subsp. hypogaea and subsp. fastigiata can be recommended based on this analysis for developing mapping populations and breeding for high yielding and resistant cultivars.

Ethylene inhibited aflatoxin biosynthesis is due to oxidative stress alleviation and related to glutathione redox state changes in Aspergillus flavus

The effect of 2-chloroethyl phosphoric acid (CEPA) on aflatoxin biosynthesis, the expression of aflatoxin biosynthetic genes, reactive oxygen species (ROS) formation, cellular redox status, and enzymes involved in glutathione consumption and regeneration in Aspergillus flavus was investigated. The results demonstrated that CEPA dose dependently inhibited aflatoxin B(1) production. The expression of two typical genes involved in aflatoxin biosynthesis, aflR and aflD, was reduced after CEPA treatment at 7 d. Meanwhile, CEPA significantly reduced ROS production and thiobarbituric acid reactive substances (TBARS), increased the ratio of reduced glutathione (GSH) and oxidized glutathione (GSSG) at 5, 6 and 7 d. The activities of glutathione peroxidase (GPx), glutathione reductase (GR), as well as glucose-6-phosphate dehydrogenase (G6PDH) were significantly inhibited after CEPA treatment at 5, 6 and 7 d. The present study suggested that ethylene reduced aflatoxin production is due to oxidative stress alleviation of fungal cells and is related to glutathione redox state changes.

Diversity characterization and association analysis of agronomic traits in a Chinese peanut (Arachis hypogaea L.) mini-core collection

Association mapping is a powerful approach for exploring the molecular basis of phenotypic variations in plants. A peanut (Arachis hypogaea L.) mini-core collection in China comprising 298 accessions was genotyped using 109 simple sequence repeat (SSR) markers, which identified 554 SSR alleles and phenotyped for 15 agronomic traits in three different environments, exhibiting abundant genetic and phenotypic diversity within the panel. A model-based structure analysis assigned all accessions to three groups. Most of the accessions had the relative kinship of less than 0.05, indicating that there were no or weak relationships between accessions of the mini-core collection. For 15 agronomic traits in the peanut panel, generally the Q + K model exhibited the best performance to eliminate the false associated positives compared to the Q model and the general linear model-simple model. In total, 89 SSR alleles were identified to be associated with 15 agronomic traits of three environments by the Q + K model-based association analysis. Of these, eight alleles were repeatedly detected in two or three environments, and 15 alleles were commonly detected to be associated with multiple agronomic traits. Simple sequence repeat allelic effects confirmed significant differences between different genotypes of these repeatedly detected markers. Our results demonstrate the great potential of integrating the association analysis and marker-assisted breeding by utilizing the peanut mini-core collection.

Genetic Diversity and Population Structure of the Major Peanut (Arachis hypogaea L.) Cultivars Grown in China by SSR Markers

One hundred and forty-six highly polymorphic simple sequence repeat (SSR) markers were used to assess the genetic diversity and population structure of 196 peanut (Arachis Hypogaea L.) cultivars which had been extensively planted in different regions in China. These SSR markers amplified 440 polymorphic bands with an average of 2.99, and the average gene diversity index was 0.11. Eighty-six rare alleles with a frequency of less than 1% were identified in these cultivars. The largest Fst or genetic distance was found between the cultivars that adapted to the south regions and those to the north regions in China. A neighbor-joining tree of cultivars adapted to different ecological regions was constructed based on pairwise Nei’s genetic distances, which showed a significant difference between cultivars from the south and the north regions. A model-based population structure analysis divided these peanut cultivars into five subpopulations (P1a, P1b, P2, P3a and P3b). P1a and P1b included most the cultivars from the southern provinces including Guangdong, Guangxi and Fujian. P2 population consisted of the cultivars from Hubei province and parts from Shandong and Henan. P3a and P3b had cultivars from the northern provinces including Shandong, Anhui, Henan, Hebei, Jiangsu and the Yangtze River region including Sichuan province. The cluster analysis, PCoA and PCA based on the marker genotypes, revealed five distinct clusters for the entire population that were related to their germplasm regions. The results indicated that there were obvious genetic variations between cultivars from the south and the north, and there were distinct genetic differentiation among individual cultivars from the south and the north. Taken together, these results provided a molecular basis for understanding genetic diversity of Chinese peanut cultivars.

Abundant Microsatellite Diversity and Oil Content in Wild Arachis Species

The peanut (Arachis hypogaea) is an important oil crop. Breeding for high oil content is becoming increasingly important. Wild Arachis species have been reported to harbor genes for many valuable traits that may enable the improvement of cultivated Arachis hypogaea, such as resistance to pests and disease. However, only limited information is available on variation in oil content. In the present study, a collection of 72 wild Arachis accessions representing 19 species and 3 cultivated peanut accessions were genotyped using 136 genome-wide SSR markers and phenotyped for oil content over three growing seasons. The wild Arachis accessions showed abundant diversity across the 19 species. A. duranensis exhibited the highest diversity, with a Shannon-Weaver diversity index of 0.35. A total of 129 unique alleles were detected in the species studied. A. rigonii exhibited the largest number of unique alleles (75), indicating that this species is highly differentiated. AMOVA and genetic distance analyses confirmed the genetic differentiation between the wild Arachis species. The majority of SSR alleles were detected exclusively in the wild species and not in A. hypogaea, indicating that directional selection or the hitchhiking effect has played an important role in the domestication of the cultivated peanut. The 75 accessions were grouped into three clusters based on population structure and phylogenic analysis, consistent with their taxonomic sections, species and genome types. A. villosa and A. batizocoi were grouped with A. hypogaea, suggesting the close relationship between these two diploid wild species and the cultivated peanut. Considerable phenotypic variation in oil content was observed among different sections and species. Nine alleles were identified as associated with oil content based on association analysis, of these, three alleles were associated with higher oil content but were absent in the cultivated peanut. The results demonstrated that there is great potential to increase the oil content in A. hypogaea by using the wild Arachis germplasm.

Genomic affinities of Arachis genus and interspecific hybrids were revealed by SRAP markers.

The Arachis genus is native to South America, and contains 70–80 described species assembled into nine sections. A better understanding of the level of speciation and taxonomic relationships is a prerequisite to the effective use of Arachis species in peanut breeding programs. Forty-eight genotypes representing 19 species in 6 sections were evaluated to assay the genetic variability within and among species, and 10 recombinant lines and those parents were identified with introgression of Arachis species chromosome segments into A.hypogaea genome using SRAP markers. Sixty of sixty-four SRAP primers tested were selected for DNA amplification reactions. A dendrogram and principal component analysis were constructed based on 353 SRAP polymorphic bands of the accessions. The number of scored polymorphic bands per each primer combination varied from 1 to 25 with an average of 5.9 per reaction. Estimates of genetic distance among the 48 accessions Arachis species ranged from 0.11 to 0.76. A-genome accessions 475845 (A. duranensis), and 331197 (A. villosa) were most closely associated to A.hypogaea. The first two PCAs accounted for 77.74% (62.02 and 15.72%) of the total variation observed and separated the different genomic groups. SRAPs also identified introgression of Arachis species chromosome segments into A.hypogaea. genome with 10 recombinant lines and those parents. The present results indicated that SRAPs can be used to determine the genetic relationships among species of the different sections of the genus Arachis and to identify introgression of Arachis genus chromosome segments into A.hypogaea genome.

Genomic Tools in Groundnut Breeding Program: Status and Perspectives

Groundnut, a nutrient-rich food legume, is cultivated world over. It is valued for its good quality cooking oil, energy and protein rich food, and nutrient-rich fodder. Globally, groundnut improvement programs have developed varieties to meet the preferences of farmers, traders, processors, and consumers. Enhanced yield, tolerance to biotic and abiotic stresses and quality parameters have been the target traits. Spurt in genetic information of groundnut was facilitated by development of molecular markers, genetic, and physical maps, generation of expressed sequence tags (EST), discovery of genes, and identification of quantitative trait loci (QTL) for some important biotic and abiotic stresses and quality traits. The first groundnut variety developed using marker assisted breeding (MAB) was registered in 2003. Since then, USA, China, Japan, and India have begun to use genomic tools in routine groundnut improvement programs. Introgression lines that combine foliar fungal disease resistance and early maturity were developed using MAB. Establishment of marker-trait associations (MTA) paved way to integrate genomic tools in groundnut breeding for accelerated genetic gain. Genomic Selection (GS) tools are employed to improve drought tolerance and pod yield, governed by several minor effect QTLs. Draft genome sequence and low cost genotyping tools such as genotyping by sequencing (GBS) are expected to accelerate use of genomic tools to enhance genetic gains for target traits in groundnut.

Recent Advances in Molecular Genetic Linkage Maps of Cultivated Peanut

The competitiveness of peanuts in domestic and global markets has been threatened by losses in productivity and quality that are attributed to diseases, pests, environmental stresses and allergy or food safety issues. Narrow genetic diversity and a deficiency of polymorphic DNA markers severely hindered construction of dense genetic maps and quantitative trait loci (QTL) mapping in order to deploy linked markers in marker-assisted peanut improvement. The U.S. Peanut Genome Initiative (PGI) was launched in 2004, and expanded to a global effort in 2006 to address these issues through coordination of international efforts in genome research beginning with molecular marker development and improvement of map resolution and coverage. Ultimately, a peanut genome sequencing project was launched in 2012 by the Peanut Genome Consortium (PGC). We reviewed the progress for accelerated development of peanut genomic resources in peanut, such as generation of expressed sequenced tags (ESTs) (252,832 ESTs as December 2012 in the public NCBI EST database), development of molecular markers (over 15,518 SSRs), and construction of peanut genetic linkage maps, in particular for cultivated peanut. Several consensus genetic maps have been constructed, and there are examples of recent international efforts to develop high density maps. An international reference consensus genetic map was developed recently with 897 marker loci based on 11 published mapping populations. Furthermore, a high-density integrated consensus map of cultivated peanut and wild diploid relatives also has been developed, which was enriched further with 3693 marker loci on a single map by adding information from five new genetic mapping populations to the published reference consensus map

Deep sequencing analysis of transcriptomes in Aspergillus flavus in response to resveratrol

BackgroundResveratrol has been reported as a natural phytoalexin that inhibits infection or the growth of certain fungi including Aspergillus flavus. Our previous research revealed that aflatoxin production in A. flavus was reduced in medium with resveratrol. To understand the molecular mechanism of the A. flavus response to resveratrol treatment, the high-throughput paired-end RNA-Seq was applied to analyze the transcriptomic profiles of A. flavus.ResultsIn total, 366 and 87 genes of A. flavus were significantly up- and down- regulated, respectively, when the fungus was treated with resveratrol. Gene Ontology (GO) functional enrichment analysis revealed that 48 significantly differentially expressed genes were involved in 6 different terms. Most genes in the aflatoxin biosynthetic pathway genes cluster (#54) did not show a significant change when A. flavus was treated with resveratrol, but 23 of the 30 genes in the #54 cluster were down-regulated. The transcription of aflA and aflB was significantly suppressed under resveratrol treatment, resulting in an insufficient amount of the starter unit hexanoate for aflatoxin biosynthesis. In addition, resveratrol significantly increased the activity of antioxidative enzymes that destroy radicals, leading to decreased aflatoxin production. Moreover, stuA, fluG, flbC, and others genes involved in mycelial and conidial development were down-regulated, which disrupted the cell’s orderly differentiation and blocked conidia formation and mycelia development. The transcripts of laeA and veA were slightly inhibited by resveratrol, which may partly decrease aflatoxin production and depress conidia formation.ConclusionsResveratrol can affect the expression of A. flavus genes that are related to developmental and secondary metabolic processes, resulting in decreased aflatoxin production and conidia formation and could also cause abnormal mycelia development. These results provide insight into the transcriptome of A. flavus in response to resveratrol and a new clew for further study in regulation of aflatoxin biosynthesis in A. flavus.