Construction of High-Density Genetic Maps and Detection of QTLs Associated With Huanglongbing Tolerance in Citrus

Huanglongbing (HLB), or citrus greening, is the most devastating disease in citrus worldwide. Commercial citrus varieties including sweet orange (Citrus sinensis) are highly susceptible to HLB, and trifoliate orange (Poncirus trifoliata, a close Citrus relative) is widely considered resistant or highly tolerant to HLB. In this study, an intergeneric F1 population of sweet orange and trifoliate orange was genotyped by Genotyping-by-Sequencing, and high-density SNP-based genetic maps were constructed separately for trifoliate orange and sweet orange. The two genetic maps exhibited high synteny and high coverage of the citrus genome. Progenies of the F1 population and their parents were planted in a replicated field trial, exposed to intense HLB pressure for 3 years, and then evaluated for susceptibility to HLB over 2 years. The F1 population exhibited a wide range in severity of HLB foliar symptom and canopy damage. Genome-wide QTL analysis based on the phenotypic data of foliar symptom and canopy damage in 2 years identified three clusters of repeatable QTLs in trifoliate orange linkage groups LG-t6, LG-t8 and LG-t9. Co-localization of QTLs for two traits was observed within all three regions. Additionally, one cluster of QTLs in sweet orange (linkage group LG-s7) was also detected. The majority of the identified QTLs each explained 18–30% of the phenotypic variation, indicating their major role in determining HLB responses. These results show, for the first time, a quantitative genetic nature yet the presence of major loci for the HLB tolerance in trifoliate orange. The results suggest that sweet orange also contains useful genetic factor(s) for improving HLB tolerance in commercial citrus varieties. Findings from this study should be very valuable and timely to researchers worldwide as they are hastily searching for genetic solutions to the devastating HLB crisis through breeding, genetic engineering, or genome editing

Development and implementation of high-throughput SNP genotyping in barley

<p>Abstract</p> <p>Background</p> <p>High density genetic maps of plants have, nearly without exception, made use of marker datasets containing missing or questionable genotype calls derived from a variety of genic and non-genic or anonymous markers, and been presented as a single linear order of genetic loci for each linkage group. The consequences of missing or erroneous data include falsely separated markers, expansion of cM distances and incorrect marker order. These imperfections are amplified in consensus maps and problematic when fine resolution is critical including comparative genome analyses and map-based cloning. Here we provide a new paradigm, a high-density consensus genetic map of barley based only on complete and error-free datasets and genic markers, represented accurately by graphs and approximately by a best-fit linear order, and supported by a readily available SNP genotyping resource.</p> <p>Results</p> <p>Approximately 22,000 SNPs were identified from barley ESTs and sequenced amplicons; 4,596 of them were tested for performance in three pilot phase Illumina GoldenGate assays. Data from three barley doubled haploid mapping populations supported the production of an initial consensus map. Over 200 germplasm selections, principally European and US breeding material, were used to estimate minor allele frequency (MAF) for each SNP. We selected 3,072 of these tested SNPs based on technical performance, map location, MAF and biological interest to fill two 1536-SNP "production" assays (BOPA1 and BOPA2), which were made available to the barley genetics community. Data were added using BOPA1 from a fourth mapping population to yield a consensus map containing 2,943 SNP loci in 975 marker bins covering a genetic distance of 1099 cM.</p> <p>Conclusion</p> <p>The unprecedented density of genic markers and marker bins enabled a high resolution comparison of the genomes of barley and rice. Low recombination in pericentric regions is evident from bins containing many more than the average number of markers, meaning that a large number of genes are recombinationally locked into the genetic centromeric regions of several barley chromosomes. Examination of US breeding germplasm illustrated the usefulness of BOPA1 and BOPA2 in that they provide excellent marker density and sensitivity for detection of minor alleles in this genetically narrow material.</p

A reference genetic map of C. clementina hort. ex Tan.; citrus evolution inferences from comparative mapping

Background: Most modern citrus cultivars have an interspecific origin. As a foundational step towards deciphering the interspecific genome structures, a reference whole genome sequence was produced by the International Citrus Genome Consortium from a haploid derived from Clementine mandarin. The availability of a saturated genetic map of Clementine was identified as an essential prerequisite to assist the whole genome sequence assembly. Clementine is believed to be a &#39;Mediterranean&#39; mandarin x sweet orange hybrid, and sweet orange likely arose from interspecific hybridizations between mandarin and pummelo gene pools. The primary goals of the present study were to establish a Clementine reference map using codominant markers, and to perform comparative mapping of pummelo, sweet orange, and Clementine. Results: Five parental genetic maps were established from three segregating populations, which were genotyped with Single Nucleotide Polymorphism (SNP), Simple Sequence Repeats (SSR) and Insertion-Deletion (Indel) markers. An initial medium density reference map (961 markers for 1084.1 cM) of the Clementine was established by combining male and female Clementine segregation data. This Clementine map was compared with two pummelo maps and a sweet orange map. The linear order of markers was highly conserved in the different species. However, significant differences in map size were observed, which suggests a variation in the recombination rates. Skewed segregations were much higher in the male than female Clementine mapping data. The mapping data confirmed that Clementine arose from hybridization between &#39;Mediterranean&#39; mandarin and sweet orange. The results identified nine recombination break points for the sweet orange gamete that contributed to the Clementine genome. Conclusions: A reference genetic map of citrus, used to facilitate the chromosome assembly of the first citrus reference genome sequence, was established. The high conservation of marker order observed at the interspecific level should allow reasonable inferences of most citrus genome sequences by mapping next-generation sequencing (NGS) data in the reference genome sequence. The genome of the haploid Clementine used to establish the citrus reference genome sequence appears to have been inherited primarily from the &#39;Mediterranean&#39; mandarin. The high frequency of skewed allelic segregations in the male Clementine data underline the probable extent of deviation from Mendelian segregation for characters controlled by heterozygous loci in male parents