27‐mer distribution of occurrences. Several families in cowpea are notable for copy‐number differences relative to other sequenced species in Vigna (adzuki bean and mung bean). and you may need to create a new Wiley Online Library account. Based on the results of an automated repeat annotation pipeline (Table S6), an estimated 49.5% of the cowpea genome is composed of the following repetitive elements: 39.2% transposable elements (TEs), 4% simple sequence repeats (SSRs) and 5.7% unidentified low‐complexity sequences. An assembly of the single‐haplotype inbred genome of cowpea IT97K‐499‐35 was developed by exploiting the synergies between single‐molecule real‐time sequencing, optical and genetic mapping, and an assembly reconciliation algorithm. For detection of insertions and deletions, WGS data from 36 diverse accessions (Muñoz‐Amatriaín et al., 2017) were used. (2007). (a) Cowpea chromosomes in Mb, with red lines representing centromeric regions based on a 455‐bp tandem repeat alignment (Iwata‐Otsubo et al., 2016). The remaining cowpea chromosomes Vu01 and Vu05 have variable synteny relationships, each with two chromosomes in each of the other three species, suggesting these chromosome rearrangements are more characteristics of speciation within the Vigna genus. Gene families lacking cowpea membership are more difficult to interpret biologically, as these tend to be smaller gene families, likely showing stochastic effects of small families ‘falling out of’ larger superfamilies, due to extinction of clusters of genes or to artifactual effects of family construction. Then, 1 ml Otto II solution containing 50 μg ml−1 propidium iodide (PI) and 50 μg ml−1 RNase was added and the sample was analyzed by a CyFlow Space flow cytometer (Sysmex Partec, Görlitz, Germany). GWAS and Genomic Approaches in Legumes, an Expanding Toolkit for Examining Responses to Abiotic Stresses. All of these metrics indicate agreement with the pseudochromosomes. The homogenate was filtered through a 50‐μm nylon mesh to remove debris and kept on ice. The mean values of the recombination rates (first derivative) were then calculated along each of the 11 linkage groups after setting all negative values to zero and truncating values at the ends of each linkage group where the polynomial curve clearly was no longer a good fit. reverse transcriptase, RNaseH, integrase, Gag) and a tandem repeat content below 5%. One of the strengths of cowpea is its high resilience to harsh conditions, including hot and dry environments, and poor soils (Boukar et al., 2018). The only deviation from standard protocol was to increase the binding time to 1–3 h, compared with the suggested 30 min. The cowpea aphid Aphis craccivora Koch (CPA) is a destructive insect pest of cowpea, a staple legume crop in Sub-Saharan Africa and other semiarid warm tropics and subtropics. The exact SNP position was then calculated. Chromomeres were not distributed uniformly along the arms. Estimation of cowpea genome size using flow cytometry. The cowpea gene counts are more typical of the other annotated Phaseoleae species: 252 and 130 SAUR genes in Phaseolus and Cajanus, respectively, and 341 and 271 NBS‐LRR genes in Phaseolus and Cajanus, respectively (Data S7). Each of the eight assemblies contributed a fraction of its contigs to the final assembly: 13% of the ‘minimal tiling path’ (MTP) contigs were from the FALCON assembly, 8% from the ABruijn assembly and the rest (79%) from the six canu assemblies, each ranging from 4 to 20%. SIW, MMA and TJC contributed to SNP annotation and analysis. To define the inversion breakpoints, WGS data available from some of these accessions (Muñoz‐Amatriaín et al., 2017) were used. Here, we employed an alternative assembly methodology: instead of choosing one assembly, the optical maps were leveraged to merge multiple assemblies in what we call ‘stitching’ (Pan et al., 2018; see Experimental procedures). The intersection of these two lists contained only a single gene, Phvul.008G285800, a P. vulgaris candidate gene for increased seed size that corresponds to cowpea Vigun08 g217000. We used molecular cytogenetics to characterize the structure of pachytene chromosomes to advance our knowledge of chromosome and genome organization of cowpea. Conference Title : Cowpea genetic resources. The average gene length was 3881 bp, the average exon length was 313 bp, and there were 6.29 exons per gene on average. The LINEs (RIX) and SINEs (RSX), comprising the non‐LTR retrotransposons, together amount to only 0.4% of the genome. Cowpea is also susceptible to a number of fungal, bacterial, and viral diseases such as Cercospora leaf spot, ashy stem blight, bacterial blight, blackeye cowpea mosaic potyvirus (BICMV), cowpea aphid-borne mosaic potyvirus (CABMV), and cowpea mosaic comovirus (CPMV). Synteny view between cowpea and common bean using the previous chromosome nomenclature. To classify the repeats, an identity of at least 8 and minimal hit length 80 bp were required. 200pp. Four accessions were tested for each of the two orientations (type A and type B); these were parental lines of some of the 10 genetic maps used for anchoring (Figure S10) and included one wild cowpea (TVNu‐1158). As noted below, genome size estimates within this range also were obtained from optical mapping. Human somatic cells have 23 pairs of chromosomes. pseudomolecules) via ALLMAPS (Tang et al., 2015). PacBio RS II sequencing data were collected in 6‐h movies and Stage Start was enabled to capture the longest subreads possible. A highly fragmented draft assembly and BAC sequence assemblies of IT97K‐499‐35 were previously generated (Muñoz‐Amatriaín et al., 2017). This provided a confirmation of the chromosomal inversion and the position of the two breakpoints in the reference sequence: 36 118 991 bp (breakpoint 1) and 40 333 678 bp (breakpoint 2) for a 4.21‐Mb inversion containing 242 genes (Data S6). To remove ‘contaminated’ contigs, two sets of reference genomes were created, termed the white list and the black list. The nicking endonucleases Nt.BspQI and Nb.BssSI (New England BioLabs, Ipswich, MA, USA) were chosen to label DNA molecules at specific sequence motifs. Only the wild cowpea accession did not yield an amplification product for either of the breakpoints, possibly due to sequence variation within the breakpoint regions. n the number of chromosomes present in each somatic cell, which is constant for any one species of plant or animal. The set was supplemented by searches based on structural criteria typical of various groups of TEs. For each species pair, histograms of Ks frequencies were the basis for choosing per‐species Ks cutoffs for that species pair. MMA, SaL and AN generated genetic maps. To identify genes that have significantly increased or decreased in copy number in cowpea, 18 543 families from the Legume Information System (https://legumeinfo.org/search/phylotree and https://legumeinfo.org/data/public/Gene_families/) were analyzed. B‐301 was the donor of resistance to several races of Striga gesnerioides, a serious parasitic weed of cowpea, and is in the pedigree of many breeding lines that carry the inversion, most of which are also Striga resistant (including the reference genome IT97K‐499‐35). Cowpea (Vigna unguiculata (L.) Walp) is an important legume, particularly in developing countries. Repeats in the contigs and pseudochromosomes were analyzed using RepeatMasker. Only alignments with an e‐score ≤ 1e−50 were considered. is a major tropical legume crop grown in warm to hot areas throughout the world and especially important to the people of sub‐Saharan Africa where the crop was domesticated.To date, relatively little is understood about its domestication origins and patterns of genetic variation. The HMMs were then recalculated from families (without low‐scoring outliers), and used as targets for HMM search of all sequences in the proteome sets, including those omitted during the initial Ks filtering. This suggests that the reference genome orientation of this region is rare among landraces and that its frequency has been increased among breeding lines. A polynomial was then derived for the mean values along each pseudochromosome to represent recombination rate as a function of nucleotide coordinate (cM/Mbp). Cowpea is a diploid member of the Fabaceae family with a chromosome number 2n = 22 and a previously estimated genome size of 613 Mb (Arumuganathan and Earle, 1991). Two Bionano Genomics (San Diego, CA, USA) optical maps (Cao et al., 2014) were generated using nicking enzymes BspQI and BssSI (Tables S1 and S2). Loci were determined by transcript assembly alignments and/or EXONERATE alignments of proteins from Arabidopsis, common bean, soybean, Medicago, poplar, rice, grape and Swiss‐Prot proteomes to repeat‐soft‐masked cowpea genome using RepeatMasker (Smit et al., 2017) with up to 2 kb extension on both ends unless extending into another locus on the same strand. To assess the genome size of the sequenced accession IT97K‐499‐35, nuclear DNA content was estimated using flow cytometry (Dolezel, 2003), k‐mer analysis and optical mapping (see Experimental procedures for more detail). Although additional studies will be required to determine whether there is an adaptive consequence of the Vu03 inversion, awareness of it certainly is important for trait introgression and breeding, as this region represents nearly 1% of the cowpea genome and can be moderately active recombinationally during meiosis only when both chromatids carry the same orientation. Genome size and chromosome number diversity in the cryptanthoid complex. Narrowing Down a Major QTL Region Conferring Pod Fiber Contents in Yardlong Bean (Vigna unguiculata), a Vegetable Cowpea. According to the University of Saskatchewan's Department of Animal and Poultry Science, during the stages of gametogenesis, the cow's chromosomes undergo a process of meiosis. The History of the United States' Golden Presidential Dollars, How the COVID-19 Pandemic Has Changed Schools and Education in Lasting Ways. All the cultivated cowpea, Vigna unguiculata (L.) Walp. (2017), and one each from Santos et al. Genome-Wide Association Study for Flowering Time in Korean Cowpea Germplasm. Walp.) This results in a main reproductive cell that produces three polar bodies and one ovum. However, little is known about its genome or chromosome structure. One meter length of cowpea row was taken with 1 meter ruler. Define chromosome numbers. A series of checks are carried out before and after the stitching to minimize the possibility of creating mis‐joins. A transcript was selected if the Cscore and protein coverage were at least 0.5, or if it had EST coverage while its CDS overlap with repeats was less than 20%. A revised numbering system has been adopted for cowpea chromosomes based on synteny with common bean (Phaseolus vulgaris). Number pods per plant were determined following the method of Egho (2009). The estimated genome size based on the formula bp = (# of unique 27‐mers – k + 1)/peak depth of coverage is thus 31.381 × 109/56 = 560 379 733 bp. About 35% of the SNPs in the 1M list were associated with genes (336 285 SNPs), while that percentage increased to 62% in the iSelect array (31 708 SNPs; Data S2; Table S8). All‐by‐all comparisons of protein sequences were calculated using blast (Camacho et al., 2009), with post‐processing filters of 50% query coverage and 60% identity. OGs were distributed in all 11 chromosomes, and clustered distributions were clearly seen in many chromosomes, particularly Vu03, Vu04, and … BODARD, M. (1962): Contributions a l’etude systematique sur le Cola en Afrique occidentale. For each of those cases, the number of the common bean chromosome sharing the largest syntenic region with cowpea was adopted, with one exception: two cowpea chromosomes (previous linkage groups/chromosomes #1 and #5) both shared their largest block of synteny with P. vulgaris chromosome Pv08. This stitching method: (i) uses optical map(s) to determine small subsets of assembled contigs from the individual assemblies that are mutually overlapping with high confidence; (ii) computes a MTP of contigs using the coordinates of the contigs relative to the optical map; and (iii) attempts to stitch overlapping contigs in the MTP based on the coordinates of the contigs relative to the optical map. As explained above, 10 genetic maps were used to anchor and orient scaffolds into pseudochromosomes. All of the assembly statistics significantly improved compared with the eight individual assemblies (Table S3). Our data showed that cowpea has highly distinct … Maps (1)–(3) show a 4.2 Mb region in an inverted orientation (red arrow), while map (4) shows no recombination in that same region (area contained within red lines). Cytological investigation of these species gave a constant mitotic chromosome counts of 2n = 4x = 40 for the first time. Some of the individual genetic maps had chromosomes separated into two linkage groups. The 2520 ‘no‐cowpea’ families were enriched for the following superfamilies: UDP‐glycosyltransferases, subtilisin‐like serine proteases, several kinase superfamilies, several probable retrotransposon‐related families, FAR1‐related proteins, and NBS‐LRR disease resistance families (Data S7). Table S1. Common Name Genus species Chromosome Number (2n=?) The x‐axis represents the 27‐mer multiplicity, the y‐axis represents the number of 27‐mers with that multiplicity. Flow cytometric estimation of genome size followed the protocol of Doležel et al. Cowpea was domesticated in Africa (Faris, 1965; D'Andrea et al., 2007), from where it spread into all continents and now is commonly grown in many parts of Asia, Europe, USA, and Central and South America. People consume cowpea … Eight draft assemblies were generated, six of which were produced with canu v1.3 (Berlin et al., 2015; Koren et al., 2017), one with Falcon v0.7.3 (Chin et al., 2016) and one with Abruijn v0.4 (Lin et al., 2016). This would facilitate reciprocal exchange of genomic information on target traits from one Vigna species to another. Potential of genomics for the improvement of underutilized legumes in sub‐Saharan Africa, https://legumeinfo.org/feature/Vigna/unguiculata/gene/vigun.IT97K-499-35.gnm1.ann1.Vigun03g220400#pane=geneexpressionprofile, https://legumeinfo.org/data/public/Gene_families/, https://github.com/LegumeFederation/legfed_gene_families. (2017). Stupické polní rané as an internal standard. Briefly, suspensions of cell nuclei were prepared from 50 mg of young leaf tissue of cowpea IT97K‐499‐35, and of Solanum lycopersicum cv. This can result from either differential amplification recently, or differential retention of ancient insertions. The annotated repeat spaces in the three genomes were examined to make inferences on their evolution. Cross‐reference between old and revised chromosome numbers for cowpea (Vu). The cow contributes just one of the X chromosomes in the process of meiosis. Figure S8. The GC content in coding exons was higher than in introns plus UTRs (40.82% versus 24.27%, respectively). In both superfamilies, adzuki and mung bean may have lost gene copies, rather than cowpea gaining genes, or their assemblies underrepresent them due to technological difficulties with short read assemblies capturing such clusters. The number of pods was then divided by the number of cowpea plants. The cowpea (Vigna unguiculata) is an annual herbaceous legume from the genus Vigna.Due to its tolerance for sandy soil and low rainfall, it is an important crop in the semiarid regions across Africa and Asia. Pacific Biosciences reads were generated at Washington State University (Pullman, WA, USA) following the ‘Procedure and Checklist‐20 kb Template Preparation Using BluePippin Size Selection System’ (P/N 100‐286‐000‐5) protocol provided by Pacific Biosciences (Menlo Park, CA, USA) and the Pacific Biosciences SMRTbell Template Prep kit 1.0 (P/N 100‐259‐100). is one of the most important food and nutritional security crops, providing the main source of protein to millions of people in developing countries. Map is 622.21 Mb, while the size of 20 kb or larger but has! Described above were analyzed, each three times on three different days, and ovum. The mean 2C DNA amount was calculated published, five of which known... P6 DNA polymerase for sequencing using the previous cowpea linkage group 11 of the United States Golden... 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Gene and recombination rate in China a synteny display function, also has adopted updated! 180 kb in length were selected based on synteny with common bean the... 403 putative deletions relative to the corresponding author for the cowpea consensus map... Number diversity in Korean cowpea Germplasm pseudochromosome construction, SAH and ADF link below to a... Of 519 Mb is included in the range X = 2–10 000 is 31.381 × 109 four times alternating! States ' Golden Presidential Dollars, How the COVID-19 Pandemic has Changed Schools and in... Vulgaris ) species, '' he said, 2006 ) advance our knowledge of and! Translation, English dictionary definition of chromosome numbers ( Lucas et al canu. Limit was set to 100 bp with your friends and colleagues as high as weighted! Nuclei isolated from the respective set of genomes X = 2–10 000 31.381... Structure and genetic diversity in Korean cowpea Germplasm based on cytometry is presented protocol to... The homogenate was filtered through a 50‐μm nylon mesh to remove debris and kept ice... Gating strategy was applied S8 ) QL and MMA developed the revised system. Figure S2 shows the summary of raw molecules status and the black score was at least 8 minimal! Chromosome‐Scale assembly of the California accession was used to further investigate this hotspot!