Cassava (Manihot esculenta) provides calories and nutrition for more than half a billion people. It was domesticated by native Amazonian peoples through cultivation of the wild progenitor M. esculenta ssp. flabellifolia and is now grown in tropical regions worldwide. Here we provide a high-quality genome assembly for cassava with improved contiguity, linkage, and completeness; almost 97% of genes are anchored to chromosomes. We find that paleotetraploidy in cassava is shared with the related rubber tree Hevea, providing a resource for comparative studies. We also sequence a global collection of 58 Manihot accessions, including cultivated and wild cassava accessions and related species such as Ceará or India rubber (M. glaziovii), and genotype 268 African cassava varieties. We find widespread interspecific admixture, and detect the genetic signature of past cassava breeding programs. As a clonally propagated crop, cassava is especially vulnerable to pathogens and abiotic stresses. This genomic resource will inform future genome-enabled breeding efforts to improve this staple crop. 13 International Institute of Tropical Agriculture (IITA), Nairobi, Kenya. 14 Dow AgroSciences, Indianapolis, Indiana, USA. 15 Molecular Genetics Unit, Okinawa Institute of Science and Technology Graduate University, Onna, Japan. 16 In this report we use "cassava" to refer to cultivated and/or domesticated varieties of M. esculenta, and the shorthand M. esc. flabellifolia for wild accessions 3 . We also shotgun-sequenced five Manihot accessions related to cassava, including three from the wild species M. glaziovii Muell. Arg., one named M. pseudoglaziovii Pax & K. Hoffman, and "tree" cassava, a suspected hybrid sometimes called M. catingea Ule 12,18 . The Ceará or India rubber tree species M. glaziovii, also domesticated in South America, was imported to East Africa in the early twentieth century. It is interfertile with cassava and has been used in African breeding programs to exploit the natural resistance of M. glaziovii to cassava pathogens 18 . To analyze genetic variation present in African varieties, we also characterized 268 cultivars of cassava using reduced representation genotypingby-sequencing (GBS) 19 (Table 2). RESULTS Chromosome structureTo produce a high-quality chromosome-scale reference genome for cassava, we augmented our earlier draft sequence 20 of the reference genotype AM560-2 with additional whole genome shotgun sequencing and mate pair data, fosmid-end sequences, and a paired-end library developed using proximity ligation of in vitro reconstituted chromatin 21 (Methods and Supplementary Note 1). AM560-2 is an S3 line bred at Centro Internacional de Agricultura Tropical (CIAT) from MCOL1505 (also known as Manihoica P-12 (ref. 22). Compared with the previous draft 23 , the contiguity of our new shotgun assembly has more than doubled (N50 length 27.7 kb vs. 11.5 kb), and an additional 135 Mb is anchored to chromosomes 23 (Supplementary Note 1). To organize the sequence into chromosomes we integrated the shotgun ...

Cassava mosaic disease (CMD), caused by different species of cassava mosaic geminiviruses (CMGs), is the most important disease of cassava in Africa and the Indian sub-continent. The cultivated cassava species is protected from CMD by polygenic resistance introgressed from the wild species Manihot glaziovii and a dominant monogenic type of resistance, named CMD2, discovered in African landraces. The ability of the monogenic resistance to confer high levels of resistance in different genetic backgrounds has led recently to its extensive usage in breeding across Africa as well as pre-emptive breeding in Latin America. However, most of the landraces carrying the monogenic resistance are morphologically very similar and come from a geographically restricted area of West Africa, raising the possibility that the diversity of the single-gene resistance could be very limited, or even located at a single locus. Several mapping studies, employing bulk segregant analysis, in different genetic backgrounds have reported additional molecular markers linked to supposedly new resistance genes. However, it is not possible to tell if these are indeed new genes in the absence adequate genetic map framework or allelism tests. To address this important question, a high-density single nucleotide polymorphism (SNP) map of cassava was developed through genotyping-by-sequencing a bi-parental mapping population (N=180) that segregates for the dominant monogenic resistance to CMD. Virus screening using PCR showed that CMD symptoms and presence of virus were strongly correlated (r=0.98). Genome-wide scan and high-resolution composite interval mapping using 6756 SNPs uncovered a single locus with large effect (R(2)=0.74). Projection of the previously published resistance-linked microsatellite markers showed that they co-occurred in the same chromosomal location surrounding the presently mapped resistance locus. Moreover, their relative distance to the mapped resistance locus correlated with the reported degree of linkage with the resistance phenotype. Cluster analysis of the landraces first shown to have this type of resistance revealed that they are very closely related, if not identical. These findings suggest that there is a single source of monogenic resistance in the crop's genepool tracing back to a common ancestral clone. In the absence of further resistance diversification, the long-term effectiveness of the single gene resistance is known to be precarious, given the potential to be overcome by CMGs due to their fast-paced evolutionary rate. However, combining the quantitative with the qualitative type of resistance may ensure that this resistance gene continues to offer protection to cassava, a crop that is depended upon by millions of people in Africa against the devastating onslaught of CMGs.

Cassava (Manihot esculenta Crantz) is a crucial, under-researched crop feeding millions worldwide, especially in Africa. Cassava mosaic disease (CMD) has plagued production in Africa for over a century. Biparental mapping studies suggest primarily a single major gene mediates resistance. To investigate this genetic architecture, we conducted the first genome-wide association mapping study in cassava with up to 6128 genotyping-by-sequenced African breeding lines and 42,113 reference genome-mapped single-nucleotide polymorphism (SNP) markers. We found a single region on chromosome 8 that accounts for 30 to 66% of genetic resistance in the African cassava germplasm. Thirteen additional regions with small effects were also identified. Further dissection of the major quantitative trait locus (QTL) on chromosome 8 revealed the presence of two possibly epistatic loci and/or multiple resistance alleles, which may account for the difference between moderate and strong disease resistances in the germplasm. Search of potential candidate genes in the major QTL region identified two peroxidases and one thioredoxin. Finally, we found genomic prediction accuracy of 0.53 to 0.58 suggesting that genomic selection (GS) will be effective both for improving resistance in breeding populations and identifying highly resistant clones as varieties.

T he revolution in sequencing technologies has enabled fast and relatively inexpensive genome information (Metzker, 2010 Abbreviations: AMMI, additive main effect and multiplicative interaction; a top10 , mean relatedness of the top10 individuals in the validation set to those in the training set; AYT, advanced yield trial; BLUE, best linear unbiased estimator; BLUP, best linear unbiased predictor; CMD, cassava mosaic disease; CMDI, cassava mosaic disease incidence; CV-CR, cross-validation close relatives; CV-GE, crossvalidation genotype × environment; CV-Random, random crossvalidation; CV-Random_Half, cross-validation scheme in which a randomly chosen half of the observations are used; CV-noCR, cross-validation no close relatives; DM, root dry matter content; GS, genomic selection; G×E, genotype × environment; GBS, genotyping by sequencing; MAS, marker-assisted selection; MCBBI, mean cassava bacterial blight incidence; PYT, preliminary yield trial; RCBD, randomized complete-block design; RKHS, reproducing kernel Hilbert spaces; SNP, single nucleotide polymorphism; top10, 10 most closely related individuals; UYT, uniform yield trial.

Several inbred lines of acetolactate synthase (ALS)-inhibiting herbicide-resistant (ALS-R) Palmer amaranth and ALS-susceptible (ALS-S) common waterhemp were developed in the greenhouse. Interspecific hybrids were obtained by allowing several ALS-S common waterhemp females to be pollinated only by ALS-R Palmer amaranth in a growth chamber. Putative hybrid progeny were treated with an ALS-inhibiting herbicide, and the hybrid nature verified using a polymorphism found in the parental ALS gene. Polymerase chain reaction (PCR) was used to amplify a region of the ALS gene in both parental plants and putative hybrids. Restriction enzyme digestion of the ALS-R Palmer amaranth PCR fragment resulted in two smaller fragments, whereas the PCR fragment in the ALS-S common waterhemp was not cut. Restriction digestion of the putative hybrid PCR fragment showed a combination of ALS-R Palmer amaranth double fragments and an ALS-S common waterhemp single fragment. Approximately 4 million flowers were present on 11 common waterhemp females and produced about 44,000 seeds that appeared viable. From the approximately 3,500 putative hybrid seedlings that were screened, 35 were confirmed as hybrids using herbicide resistance as a phenotypic and molecular marker. The data collected here verify that interspecific hybridization does occur between these two species, and even at a low rate, it could contribute to the rapid spread of ALS resistance in these species.