Telomere-to-telomere and haplotype-resolved genome of the kiwifruit Actinidia eriantha.

Actinidia eriantha is a characteristic fruit tree featuring with great potential for its abundant vitamin C and strong disease resistance. It has been used in a wide range of breeding programs and functional genomics studies. Previously published genome assemblies of A. eriantha are quite fragmented and not highly contiguous. Using multiple sequencing strategies, we get the haplotype-resolved and gap-free genomes of an elite breeding line "Midao 31" (MD), termed MDHAPA and MDHAPB. The new assemblies anchored to 29 pseudochromosome pairs with a length of 619.3 Mb and 611.7 Mb, as well as resolved 27 and 28 gap-close chromosomes in a telomere-to-telomere (T2T) manner. Based on the haplotype-resolved genome, we found that most alleles experienced purifying selection and coordinately expressed. Owing to the high continuity of assemblies, we defined the centromeric regions of A. eriantha, and identified the major repeating monomer, which is designated as Ae-CEN153. This resource lays a solid foundation for further functional genomics study and horticultural traits improvement in kiwifruit.

quarTeT: a telomere-to-telomere toolkit for gap-free genome assembly and centromeric repeat identification.

A high-quality genome is the basis for studies on functional, evolutionary, and comparative genomics. The majority of attention has been paid to the solution of complex chromosome structures and highly repetitive sequences, along with the emergence of a new 'telomere-to-telomere (T2T) assembly' era. However, the bioinformatic tools for the automatic construction and/or characterization of T2T genome are limited. Here, we developed a user-friendly web toolkit, quarTeT, which currently includes four modules: AssemblyMapper, GapFiller, TeloExplorer, and CentroMiner. First, AssemblyMapper is designed to assemble phased contigs into the chromosome-level genome by referring to a closely related genome. Then, GapFiller would endeavor to fill all unclosed gaps in a given genome with the aid of additional ultra-long sequences. Finally, TeloExplorer and CentroMiner are applied to identify candidate telomere and centromere as well as their localizations on each chromosome. These four modules can be used alone or in combination with each other for T2T genome assembly and characterization. As a case study, by adopting the entire modular functions of quarTeT, we have achieved the Actinidia chinensis genome assembly that is of a quality comparable to the reported genome Hongyang v4.0, which was assembled with the addition of manual handling. Further evaluation of CentroMiner by searching centromeres in Arabidopsis thaliana and Oryza sativa genomes showed that quarTeT is capable of identifying all the centromeric regions that have been previously detected by experimental methods. Collectively, quarTeT is an efficient toolkit for studies of large-scale T2T genomes and can be accessed at http://www.atcgn.com:8080/quarTeT/home.html without registration.

Telomere-to-telomere and gap-free reference genome assembly of the kiwifruit Actinidia chinensis.

Kiwifruit is an economically and nutritionally important fruit crop with extremely high contents of vitamin C. However, the previously released versions of kiwifruit genomes all have a mass of unanchored or missing regions. Here, we report a highly continuous and completely gap-free reference genome of Actinidia chinensis cv. 'Hongyang', named Hongyang v4.0, which is the first to achieve two de novo haploid-resolved haplotypes, HY4P and HY4A. HY4P and HY4A have a total length of 606.1 and 599.6 Mb, respectively, with almost the entire telomeres and centromeres assembled in each haplotype. In comparison with Hongyang v3.0, the integrity and contiguity of Hongyang v4.0 is markedly improved by filling all unclosed gaps and correcting some misoriented regions, resulting in ~38.6-39.5 Mb extra sequences, which might affect 4263 and 4244 protein-coding genes in HY4P and HY4A, respectively. Furthermore, our gap-free genome assembly provides the first clue for inspecting the structure and function of centromeres. Globally, centromeric regions are characterized by higher-order repeats that mainly consist of a 153-bp conserved centromere-specific monomer (Ach-CEN153) with different copy numbers among chromosomes. Functional enrichment analysis of the genes located within centromeric regions demonstrates that chromosome centromeres may not only play physical roles for linking a pair of sister chromatids, but also have genetic features for participation in the regulation of cell division. The availability of the telomere-to-telomere and gap-free Hongyang v4.0 reference genome lays a solid foundation not only for illustrating genome structure and functional genomics studies but also for facilitating kiwifruit breeding and improvement.