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

Abstract

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.

Acknowledgements

This work was supported by funds from the National Natural Science Foundation of China (31972474, 31471157) and the Open Fund of State Key Laboratory of Tea Plant Biology and Utilization (SKLTOF20150103).

Author contributions

Y.Liu, J.Y., and Y.Y. conceived the ideas for this paper. X.W., S.Cao and J.Y. provided good guidance for algorithm design. Y.Lin designed the main program. Q.C., Y.Wu, F.Z., S.Z., and Y. Wang tested the program and provided critical advice. C.Y., X.L., S.Chen, and Y.Y. developed and implemented the web server. Q.C., Y.Wu, F.Z., R.P., and J.Y. illustrated images used in web pages and gave good advice on drawing figures. Y.Lin drafted the first manuscript. Y.Liu and J.Y.edited the manuscript. All authors read and approved the final manuscript.

Data availability

The source code of the quarTeT toolkit command-line program is publicly available on GitHub (https://github.com/aaranyue/ quarTeT). The dataset of HY4Q is available on the download page of the quarTeT website (http://atcgn.com:8080/quarTeT/ download.html).

Conflict of interest statement

None declared.

References

[1] Mehrotra S, Goyal V . Repetitive sequences in plant nuclear DNA: types, distribution, evolution and function. Genom Proteom Bioinf. 2014; 12: 164-71.

[2] Ahmad SF, Singchat W, Jehangir M et al. Dark matter of primate genomes: satellite DNA repeats and their evolutionary dynamics. Cell. 2020; 9: 2714.

[3] Wenger AM, Peluso P, Rowell WJ et al. Accurate circular consensus long—read sequencing improves variant detection and assembly of a human genome. Nat Biotechnol. 2019; 37: 1155-62.

[4] Jain M, Koren S, Miga KH et al. Nanopore sequencing and assembly of a human genome with ultra—long reads. Nat Biotechnol. 2018; 36: 338-45.

[5] van Dijk EL, Jaszczyszyn Y, Naquin D et al. The third revolution in sequencing technology. Trends Genet. 2018; 34: 666-81.

[6] Li H . New strategies to improve minimap2 alignment accuracy. Bioinformatics. 2021; 37: 4572-4.

[7] Cheng H, Jarvis ED, Fedrigo O et al. Haplotype—resolved assembly of diploid genomes without parental data. Nat Biotechnol. 2022; 40: 1332-5.

[8] Fu A, Zheng Y, Guo J et al. Telomere—to—telomere genome assembly of bitter melon (Momordica charantia L. var. abbreviata Ser.) reveals fruit development, composition and ripening genetic characteristics . Hortic Res. 2023; 10: uhac228.

[9] Nurk S, Koren S, Rhie A et al. The complete sequence of a human genome. Science. 2022; 376: 44-53.

[10] Li F, Xu S, Xiao Z et al. Gap—free genome assembly and comparative analysis reveal the evolution and anthocyanin accumulation mechanism of Rhodomyrtus tomentosa . Hortic Res. 2023; 10: uhad005.

[11] Shi X, Cao S, Wang X et al. The complete reference genome for grapevine (Vitis vinifera L.) genetics and breeding . Hortic Res. 2023; 10: uhad061.

[12] Zhou Y, Xiong J, Shu Z et al. The telomere—to—telomere genome of Fragaria vesca reveals the genomic evolution of Fragaria and the origin of cultivated octoploid strawberry . Hortic Res. 2023; 10: uhad027.

[13] Miga KH . Centromere studies in the era of ’telomere—to—telomere’ genomics. Exp Cell Res. 2020; 394: 112127.

[14] Ma J, Wing RA, Bennetzen JL et al. Plant centromere organization: a dynamic structure with conserved functions. Trends Genet. 2007; 23: 134—9.

[15] Mahtani MM, Willard HF . Pulsed—field gel analysis of alpha—satellite DNA at the human X chromosome centromere: high—frequency polymorphisms and array size estimate. Genomics. 1990; 7: 607-13.

[16] Nagaki K, Talbert PB, Zhong CX et al. Chromatin immunoprecipitation reveals that the 180—bp satellite repeat is the key functional DNA element of Arabidopsis thaliana centromeres . Genetics. 2003; 163: 1221-5.

[17] Benson G. Tandem repeats finder: a program to analyze DNA sequences. Nucleic Acids Res. 1999; 27: 573-80.

[18] Yue J, Chen Q, Wang Y et al. Telomere—to—telomere and gap—free reference genome assembly of the kiwifruit Actinidia chinensis . Hortic Res. 2023; 10: uhac264.

[19] Naish M, Alonge M, Wlodzimierz P et al. The genetic and epigenetic landscape of the Arabidopsis centromeres . Science. 2021; 374: eabi7489.

[20] Song J, Xie W, Wang S et al. Two gap—free reference genomes and a global view of the centromere architecture in rice. Mol Plant. 2021; 14: 1757-67.

[21] Ou S, Su W, Liao Y et al. Benchmarking transposable element annotation methods for creation of a streamlined, comprehensive pipeline. Genome Biol. 2019; 20: 275.

[22] Wu H, Ma T, Kang M et al. A high—quality Actinidia chinensis (kiwifruit) genome . Hortic Res. 2019; 6: 117.

[23] Wang Y, Dong M, Wu Y et al. Telomere—to—telomere and haplotype—resolved genome of the kiwifruit Actinidia eriantha . Mol Hortic. 2023; 3: 4.

[24] Marone MP, Singh HC, Pozniak CJ et al. A technical guide to TRI—TEX, a computational pipeline for chromosome—scale sequence assembly of plant genomes. Plant Methods. 2022; 18: 128.

[25] Alonge M, Lebeigle L, Kirsche M et al. Automated assembly scaffolding using RagTag elevates a new tomato system for high—throughput genome editing. Genome Biol. 2022; 23: 258.

[26] Dvorkina T, Bzikadze AV, Pevzner PA . The string decomposition problem and its applications to centromere analysis and assembly. Bioinformatics. 2020; 36: i93-101.

[27] Gao S, Yang X, Guo H et al. HiCAT: a tool for automatic annotation of centromere structure. Genome Biol. 2023; 24: 58.

[28] Yue J, Liu J, Tang W et al. Kiwifruit Genome Database (KGD): a comprehensive resource for kiwifruit genomics. Hortic Res. 2020; 7: 117.

[29] Marçais G, Delcher AL, Phillippy AM et al. MUMmer4: a fast and versatile genome alignment system. PLoS Comput Biol. 2018; 14: e1005944.

[30] Fu L, Niu B, Zhu Z et al. CD—HIT: accelerated for clustering the next—generation sequencing data. Bioinformatics. 2012; 28: 3150-2.

[31] Camacho C, Coulouris G, Avagyan V et al. BLAST+: architecture and applications. BMC Bioinformatics. 2009; 10: 421.

[32] Manni M, Berkeley MR, Seppey M et al. BUSCO update: novel and streamlined workflows along with broader and deeper phylogenetic coverage for scoring of eukaryotic, prokaryotic, and viral genomes. Mol Biol Evol. 2021; 38: 4647-54.

[33] Ou S, Chen J, Jiang N . Assessing genome assembly quality using the LTR Assembly Index (LAI). Nucleic Acids Res. 2018; 46: e126.

[34] Hao Z, Lv D, Ge Y et al. RIdeogram: drawing SVG graphics to visualize and map genome—wide data on the idiograms. PeerJ Comput Sci. 2020; 6: e251.