Autotetraploid Induction of Three A-Genome Wild Peanut Species, Arachis cardenasii, A. correntina, and A. diogoi.

A-genome Arachis species (AA; 2n = 2x = 20) are commonly used as secondary germplasm sources in cultivated peanut breeding, Arachis hypogaea L. (AABB; 2n = 4x = 40), for the introgression of various biotic and abiotic stress resistance genes. Genome doubling is critical to overcoming the hybridization barrier of infertility that arises from ploidy-level differences between wild germplasm and cultivated peanuts. To develop improved genome doubling methods, four trials of various concentrations of the mitotic inhibitor treatments colchicine, oryzalin, and trifluralin were tested on the seedlings and seeds of three A-genome species, A. cardenasii, A. correntina, and A. diogoi. A total of 494 seeds/seedlings were treated in the present four trials, with trials 1 to 3 including different concentrations of the three chemical treatments on seedlings, and trial 4 focusing on the treatment period of 5 mM colchicine solution treatment of seeds. A small number of tetraploids were produced from the colchicine and oryzalin gel treatments of seedlings, but all these tetraploid seedlings reverted to diploid or mixoploid states within six months of treatment. In contrast, the 6-h colchicine solution treatment of seeds showed the highest tetraploid conversion rate (6-13% of total treated seeds or 25-40% of surviving seedlings), and the tetraploid plants were repeatedly tested as stable tetraploids. In addition, visibly and statistically larger leaves and flowers were produced by the tetraploid versions of these three species compared to their diploid versions. As a result, stable tetraploid plants of each A-genome species were produced, and a 5 mM colchicine seed treatment is recommended for A-genome and related wild Arachis species genome doubling.

Initiation of genomics-assisted breeding in Virginia-type peanuts through the generation of a de novo reference genome and informative markers.

Introduction: Virginia-type peanut, Arachis hypogaea subsp. hypogaea, is the second largest market class of peanut cultivated in the United States. It is mainly used for large-seeded, in-shell products. Historically, Virginia-type peanut cultivars were developed through long-term recurrent phenotypic selection and wild species introgression projects. Contemporary genomic technologies represent a unique opportunity to revolutionize the traditional breeding pipeline. While there are genomic tools available for wild and cultivated peanuts, none are tailored specifically to applied Virginia-type cultivar development programs. Methods and respective results: Here, the first Virginia-type peanut reference genome, "Bailey II", was assembled. It has improved contiguity and reduced instances of manual curation in chromosome arms. Whole-genome sequencing and marker discovery was conducted on 66 peanut lines which resulted in 1.15 million markers. The high marker resolution achieved allowed 34 unique wild species introgression blocks to be cataloged in the A. hypogaea genome, some of which are known to confer resistance to one or more pathogens. To enable marker-assisted selection of the blocks, 111 PCR Allele Competitive Extension assays were designed. Forty thousand high quality markers were selected from the full set that are suitable for mid-density genotyping for genomic selection. Genomic data from representative advanced Virginia-type peanut lines suggests this is an appropriate base population for genomic selection. Discussion: The findings and tools produced in this research will allow for rapid genetic gain in the Virginia-type peanut population. Genomics-assisted breeding will allow swift response to changing biotic and abiotic threats, and ultimately the development of superior cultivars for public use and consumption.

Modifications to a LATE MERISTEM IDENTITY1 gene are responsible for the major leaf shapes of Upland cotton (Gossypium hirsutum L.).

Leaf shape varies spectacularly among plants. Leaves are the primary source of photoassimilate in crop plants, and understanding the genetic basis of variation in leaf morphology is critical to improving agricultural productivity. Leaf shape played a unique role in cotton improvement, as breeders have selected for entire and lobed leaf morphs resulting from a single locus, okra (l-D1), which is responsible for the major leaf shapes in cotton. The l-D1 locus is not only of agricultural importance in cotton, but through pioneering chimeric and morphometric studies, it has contributed to fundamental knowledge about leaf development. Here we show that an HD-Zip transcription factor homologous to the LATE MERISTEM IDENTITY1 (LMI1) gene of Arabidopsis is the causal gene underlying the l-D1 locus. The classical okra leaf shape allele has a 133-bp tandem duplication in the promoter, correlated with elevated expression, whereas an 8-bp deletion in the third exon of the presumed wild-type normal allele causes a frame-shifted and truncated coding sequence. Our results indicate that subokra is the ancestral leaf shape of tetraploid cotton that gave rise to the okra allele and that normal is a derived mutant allele that came to predominate and define the leaf shape of cultivated cotton. Virus-induced gene silencing (VIGS) of the LMI1-like gene in an okra variety was sufficient to induce normal leaf formation. The developmental changes in leaves conferred by this gene are associated with a photosynthetic transcriptomic signature, substantiating its use by breeders to produce a superior cotton ideotype.

Mapping and genomic targeting of the major leaf shape gene (L) in Upland cotton (Gossypium hirsutum L.).

KEY MESSAGE: A major leaf shape locus (L) was mapped with molecular markers and genomically targeted to a small region in the D-genome of cotton. By using expression analysis and candidate gene mapping, two LMI1 -like genes are identified as possible candidates for leaf shape trait in cotton. Leaf shape in cotton is an important trait that influences yield, flowering rates, disease resistance, lint trash, and the efficacy of foliar chemical application. The leaves of okra leaf cotton display a significantly enhanced lobing pattern, as well as ectopic outgrowths along the lobe margins when compared with normal leaf cotton. These phenotypes are the hallmark characteristics of mutations in various known modifiers of leaf shape that culminate in the mis/over-expression of Class I KNOX genes. To better understand the molecular and genetic processes underlying leaf shape in cotton, a normal leaf accession (PI607650) was crossed to an okra leaf breeding line (NC05AZ21). An F2 population of 236 individuals confirmed the incompletely dominant single gene nature of the okra leaf shape trait in Gossypium hirsutum L. Molecular mapping with simple sequence repeat markers localized the leaf shape gene to 5.4 cM interval in the distal region of the short arm of chromosome 15. Orthologous mapping of the closely linked markers with the sequenced diploid D-genome (Gossypium raimondii) tentatively resolved the leaf shape locus to a small genomic region. RT-PCR-based expression analysis and candidate gene mapping indicated that the okra leaf shape gene (L (o) ) in cotton might be an upstream regulator of Class I KNOX genes. The linked molecular markers and delineated genomic region in the sequenced diploid D-genome will assist in the future high-resolution mapping and map-based cloning of the leaf shape gene in cotton.