A telomere-to-telomere reference genome provides genetic insight into the pentacyclic triterpenoid biosynthesis in Chaenomeles speciosa

Chaenomeles speciosa(2n=34),a medicinal and edible plant in the Rosaceae,is commonly used in traditional Chinese medicine.To date,the lack of genomic sequence and genetic studies has impeded efforts to improve its medicinal value.Herein,we report the use of an integrative approach involving PacBio HiFi(third-generation)sequencing and Hi-C scaffolding to assemble a high-quality telomere-to-telomere genome of C.speciosa.The genome comprised 650.4 Mb with a contig N50 of 35.5 Mb.Of these,632.3 Mb were anchored to 17 pseudo-chromosomes,in which 12,4,and 1 pseudo-chromosomes were represented by a single contig,two contigs,and four contigs,respectively.Eleven pseudo-chromosomes had telomere repeats at both ends,and four had telomere repeats at a single end.Repetitive sequences accounted for 49.5%of the genome,while a total of 45515 protein-coding genes have been annotated.The genome size of C.speciosa was relatively similar to that of Malus domestica.Expanded or contracted gene families were identified and investigated for their association with different plant metabolisms or biological processes.In particular,functional annotation characterized gene families that were associated with the biosynthetic pathway of oleanolic and ursolic acids,two abundant pentacyclic triterpenoids in the fruits of C.speciosa.Taken together,this telomere-to-telomere and chromosome-level genome of C.speciosa not only provides a valuable resource to enhance understanding of the biosynthesis of medicinal compounds in tissues,but also promotes understanding of the evolution of the Rosaceae.

High Throughput Sequencing of circRNAs in Tomato Leaves Responding to Multiple Stresses of Drought and Heat

Our aim is to study the roles of a new emerging group of non-coding RNAs, circRNAs, in tomato(Solanum lycopersicum L.) plants grown at the combination of drought and heat, two of the most usual stress conditions known to frequently happen in field. Tomato seedlings from cultivar‘Jinling Meiyu’ were treated without stresses(control), at water shortage, high temperature and subjected the multiple stresses. In total, 467 circRNAs were identified with 87.82% from exon using high throughput sequencing technology. Among the circRNAs, 70 were from chr1 with the range from 23 to 49 from the other chromosomes. In detail, 156 circRNAs were shared in the four libraries, while 21, 17 and 36 circRNAs were only shown in drought, heat and multiple stresses libraries, respectively. Through a differential expression analysis, four, seven and nine circRNAs were differentially regulated in tomato at drought, heat and multiple stresses as compared with control. These circRNAs played roles on photosynthesis, starch and sucrose metabolism, RNA transport, RNA degradation, spliceosome, ribosome, etc. Our study underlined the potential role of circRNAs involved in the abiotic stress response in tomato, which might pave the way for studying biological roles of circRNAs responding to multiple stresses in plants.

Mapping of fruit apex shape related QTLs across multi-genetic backgrounds in cucumber(Cucumis sativus L.)

The shape of fruit apex is critical to appearance quality in cucumber(Cucumis sativus L.),of which the genetic basis was poorly understood,and the use of marker-assisted breeding for fruit apex improvement is not available yet.In this study,the variation of fruit apex in different cucumber ecotypes was evaluated by fruit apex angle(variation coefficient from 7.1%to 15.7%)and fruit apex index(variation coefficient from 8.8%to 22.6%).Fruit apex associated QTLs were mapped by using 145 F_(2:3) families and 155 F_(2:6) population that were derived from the cross of different ecotype cucumbers.Phenotyping of the mapping populations were conducted in four experiments in 2 years.Four majoreffect QTLs,Bfal4.1,Bfai4.1,Bfad6.1 and Bfai6.1 were consistently and reliably detected across two environments which could explain 11.6%-33.6%phenotypic variations(R^(2))in the F_(2:3) families.Three major-effect QTLs,Ofai4.1(R^(2)=13.4%-15.5%),Ofal4.1(R^(2)=10.7%-12.8%),and Ofad6.1(R^(2)=11.6%-12.4%)were stably detected in the F_(2:6) population in two experiments.Bfai4.1,Bfal4.1,Ofai4.1 and Ofal4.1 were integrated to be consensus QTL fa4.1,within which 11 candidate genes were predicted.Bfai6.1 and Bfad6.1 were integrated to be consensus QTL fa6.1.QTL interaction analysis showed that Bfai6.1 has epistatic effect with Bfai4.1.This study revealed two reliable major-effect fruit apex related QTLs across multi-genetic backgrounds and environments in cucumber.The possible candidate genes regulating the shape of fruit apex,and the relationship between cell division and fruit apex morphogenesis were discussed.

The chromosome-level Stevia genome provides insights into steviol glycoside biosynthesis

Stevia(Stevia rebaudiana Bertoni)is well known for its very sweet steviol glycosides(SGs)consisting of a common tetracyclic diterpenoid steviol backbone and a variable glycone.Steviol glycosides are 150–300 times sweeter than sucrose and are used as natural zero-calorie sweeteners.However,the most promising compounds are biosynthesized in small amounts.Based on Illumina,PacBio,and Hi-C sequencing,we constructed a chromosome-level assembly of Stevia covering 1416 Mb with a contig N50 value of 616.85 kb and a scaffold N50 value of 106.55 Mb.More than four-fifths of the Stevia genome consisted of repetitive elements.We annotated 44,143 high-confidence protein-coding genes in the high-quality genome.Genome evolution analysis suggested that Stevia and sunflower diverged~29.4 million years ago(Mya),shortly after the whole-genome duplication(WGD)event(WGD-2,~32.1 Mya)that occurred in their common ancestor.Comparative genomic analysis revealed that the expanded genes in Stevia were mainly enriched for biosynthesis of specialized metabolites,especially biosynthesis of terpenoid backbones,and for further oxidation and glycosylation of these compounds.We further identified all candidate genes involved in SG biosynthesis.Collectively,our current findings on the Stevia reference genome will be very helpful for dissecting the evolutionary history of Stevia and for discovering novel genes contributing to SG biosynthesis and other important agronomic traits in future breeding programs.

Chromosome-scale genome assembly of Cucumis hystrix-a wild species interspecifically cross-compatible with cultivated cucumber

Cucumis hystrix Chakr.(2n=2x=24)is a wild species that can hybridize with cultivated cucumber(C.sativus L.,2n=2x=14),a globally important vegetable crop.However,cucumber breeding is hindered by its narrow genetic base.Therefore,introgression from C.hystrix has been anticipated to bring a breakthrough in cucumber improvement.Here,we report the chromosome-scale assembly of C.hystrix genome(289 Mb).Scaffold N50 reached 14.1 Mb.Over 90%of the sequences were anchored onto 12 chromosomes.A total of 23,864 genes were annotated using a hybrid method.Further,we conducted a comprehensive comparative genomic analysis of cucumber,C.hystrix,and melon(C.melo L.,2n=2x=24).Whole-genome comparisons revealed that C.hystrix is phylogenetically closer to cucumber than to melon,providing a molecular basis for the success of its hybridization with cucumber.Moreover,expanded gene families of C.hystrix were significantly enriched in“defense response,”and C.hystrix harbored 104 nucleotide-binding site-encoding disease resistance gene analogs.Furthermore,121 genes were positively selected,and 12(9.9%)of these were involved in responses to biotic stimuli,which might explain the high disease resistance of C.hystrix.The alignment of whole C.hystrix genome with cucumber genome and self-alignment revealed 45,417 chromosome-specific sequences evenly distributed on C.hystrix chromosomes.Finally,we developed four cucumber-C.hystrix alien addition lines and identified the exact introgressed chromosome using molecular and cytological methods.The assembled C.hystrix genome can serve as a valuable resource for studies on Cucumis evolution and interspecific introgression breeding of cucumber.

A leaf shape mutant provides insight into PINOID Serine/Threonine Kinase function in cucumber(Cucumis sativus L.)

Optimizing leaf shape is a major challenge in efforts to develop an ideal plant type. Cucumber leaf shapes are diverse;however, the molecular regulatory mechanisms underlying leaf shape formation are unknown. In this study, we obtained a round leaf mutant(rl) from an ethyl methanesulfonate-induced mutagenesis population. Genetic analysis revealed that a single recessive gene, rl, is responsible for this mutation. A modified Mut Map analysis combined linkage mapping identified a single nucleotide polymorphism within a candidate gene,Csa1 M537400, as the mutation underlying the trait.Csa1 M537400 encodes a PINOID kinase protein involved in auxin transport. Expression of Csa1 M537400 was significantly lower in the rl mutant than in wild type, and it displayed higher levels of IAA(indole-3-acetic acid) in several tissues. Treatment of wild-type plants with an auxin transport inhibitor induced the formation of round leaves,similar to those in the rl mutant. Altered expression patterns of several auxin-related genes in the rl mutant suggest that rl plays a key role in auxin biosynthesis,transport, and response in cucumber. These findings provide insight into the molecular mechanism underlying the regulation of auxin signaling pathways in cucumber,and will be valuable in the development of an ideal plant type.

Comparative Cyto-molecular Analysis of Repetitive DNA Provides Insights into the Differential Genome Structure and Evolution of Five Cucumis Species

The genus Cucumis, includes the cucumber(2n = 14), melon(2n = 24), and other wild species, which is a good model for studying genome organization and evolution due to their variation in genome size and basic chromosome number. In this study, five Cucumis species with different geographical origins and basic chromosome numbers(i.e., C. sativus, C. hystrix, C. melo, C. anguria, and C. metuliferus) were used to identify and characterize the repetitive DNA in detail using a phylogenetic method. Comparative cyto-molecular genetic analysis of repetitive DNA was carried out using a graph-based clustering method, construction of Neighbor-Joining tree and fluorescence in situ hybridization(FISH).The results revealed that the five Cucumis species had differences in the repeat content of their genome, as well as in the composition of repetitive DNA and their genomic proportions. Three species from subgenus Melo showed a decreased tendency in both repeat types and genomic proportions, while two species from subgenus Cucumis also showed a decreased tendency in repeat types, but an increased tendency in genomic proportions. Phylogenic analysis of Cucumis Ty1/Copia, Ty3/Gypsy, and 45 S rDNA ITS regions revealed that C. sativus, C. hystrix, and C. melo were closely related species, which C. sativus and C. hystrix were closer, while C. anguria and C. metuliferus were closer to each other and further from the other three species. Differential accumulation and elimination of different repeat types divergently shaped the genomic architecture of these five Cucumis species, contributing to the genome’s evolution and diversification. Overall, these results enhance our understanding of the genomes of these five Cucumis species, and contribute to a more holistic view of genome evolution and phylogenetics of this genus.