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Ukuhlanganiswa Kwe-Genome Ye-T2T | Ukulandelana Okude Kakhulu

Ukuhlanganiswa Kwe-Genome Ye-T2T | Ukulandelana Okude Kakhulu Isithombe Esifakiwe
  • Ukuhlanganiswa Kwe-Genome Ye-T2T | Ukulandelana Okude Kakhulu
  • Ukuhlanganiswa Kwe-Genome Ye-T2T | Ukulandelana Okude Kakhulu
  • Ukuhlanganiswa Kwe-Genome Ye-T2T | Ukulandelana Okude Kakhulu

I-T2T (i-Telomere-kuya-ku-Telomere) I-Genome iyindinganiso yegolide yokuhlanganiswa kwe-genome esezingeni eliphezulu, ebhekisela ekwakhiweni kabusha kwe-genome okungenazikhala noma okungenazikhala, okusuka ku-telomere eyodwa kuya kwenye, kanye nokuphula imingcele yokuqhekeka kwe-genome evamile.

Inikwe amandla yi-core ONT ultra-long read sequencing futhi ihlanganiswe ne-multi-platform deep sequencing kanye ne-bioinformatics pipelines elungiselelwe, isixazululo se-BMKGENE T2T Genome sihlose "izifunda ezimnyama" ze-genomic ezingalawuleki kakhulu - ama-telomere (ama-nucleoprotein complexes akhethekile emaphethelweni e-chromosome eukaryotic), ama-centromere aphezulu e-organism (ama-tandem repeat array amakhulu), kanye nezinye izifunda ze-haplotype eziphindaphindayo neziyinkimbinkimbi, ebezilokhu zingaxazululeki isikhathi eside kwi-standard long-read sequencing. Ngokungafani ne-long reads evamile ehluleka ukuwela lezi zifunda futhi ibangele ukuwa kwe-sequence noma ama-chimeric contigs, i-ONT ultra-long read reads ingahlanganisa izikhala ezingahlangani kanye nezifunda eziyinkimbinkimbi. I-BMKGene izibophezele ekuletheni ama-genome e-T2T angenazikhala noma angenazikhala, asezingeni eliphezulu ezinhlobo ezahlukahlukene.

Ukwakhiwa kwe-genome ye-T2T kuvula izindawo ze-genomic eziyinkimbinkimbi ezazingafinyeleleki ngaphambili, kugcwalisa izikhala ezibalulekile zocwaningo, futhi kuhlinzeke ngedatha eyisisekelo eqinile nenembile kakhulu yezifundo ezijulile ezifaka phakathi ukuvela kwezinhlobo, ukumbiwa kwezakhi zofuzo ezisebenzayo, ukuzalanisa kwama-molecule, umuthi wokunemba kanye nolunye ucwaningo lwesayensi olusezingeni eliphezulu.

 


Imininingwane Yesevisi

I-Bioinformatics

Imiphumela yedemo

Izincwadi ezivelele

Izici Zesevisi

•Ihlinzeka ngokunemba okuphezulu, ukuhambisana okuphezulu, kanye nokuphelela okuphezulu kwe-telomere-to-telomere genome.

•Inqoba izinselele zokuhlanganisa ezindaweni eziphakathi nendawo neziphindaphindwayo kakhulu.

•Ihlaziya ukwehluka kwesakhiwo ezindaweni eziyinkimbinkimbi njenge-centromeres kanye ne-telomeres.

•Uhlola imvelaphi yama-chromosome kanye nokwakhiwa kwezakhi zofuzo, futhi uhlonza izakhi zofuzo ezibalulekile ezinquma ubulili.

Izinzuzo Zesevisi

•Ithimba lochwepheshe elisebenza isikhathi eside kakhulu elihlanganisa ukukhipha nokulandelanisa, kanye nolwazi oluphumelelayo ezinhlotsheni eziningi zezilwane.

•Ukufinyelela kokubili amapulatifomu e-PacBio kanye ne-Nanopore afundwa isikhathi eside ane-throughput ephezulu kanye namasu okulandelana aguquguqukayo.

•Ithimba elinolwazi ekuhlanganisweni kwezakhi zofuzo kanye nokuhlaziywa kwe-bioinformatics okwenziwe ngokwezifiso, elichwepheshile kumaphrojekthi ezakhi zofuzo ze-T2T.

•Amaphrojekthi e-genome aphumelele angaphezu kuka-200 kanye nezici zomthelela eziqongelelwe ezingaphezu kuka-2000.

•Izixazululo ezihlanganisiwe zokuhlola kanye ne-bioinformatic ezisekelwa amalungelo okushicilela kanye namalungelo obunikazi.

Imininingwane Yesevisi

Ucwaningo lwe-genome

Ukuhlanganiswa kwe-genome

Izinga le-Chromosome

Ukugcwaliswa kwezikhala

Isichasiselo se-Genome

I-50X Illumina NovaSeq PE150

Ama-HiFi reads angu-30X e-PacBio CCS

I-Hi-C engu-100X

Ukufundwa okude okungu-40-100X ONT Ultra

I-RNA-seq Illumina PE150 10 Gb + (ongakukhetha) Ubude obugcwele be-RNA-seq PacBio 40 Gb noma i-Nanopore 12 Gb

Izidingo Zesevisi

Ukuze uthole amasampula okulandelana kwe-Survey, i-PacBio CCS, i-Hi-C, kanye ne-transcriptome (ukuze uthole izincazelo), sicela ubheke ku-“izinga le-chromosomeizidingo zesampula yokuhlanganiswa kwe-genome".

Ku-ONT ultra-long sequencing, kunconywa amasampula ezicubu, anezindinganiso zekhwalithi ephezulu ukusekela ukukhishwa kwe-ultra-HMW DNA.

Ukuze uthole imiyalelo eningiliziwe yokulungiselela amasampula kanye nezidingo, sicela uxhumane nethimba lethu lokuthengisa ukuze uthole ikhambi elenziwe ngokwezifiso ngokusekelwe ohlotsheni.

Ukugeleza Komsebenzi Wesevisi

Isampula ye-QC

Umklamo wokuhlola

ukulethwa kwesampula

Ukulethwa kwesampula

Ukuhlolwa kwe-pilot

Ukukhishwa kwe-DNA

Ukulungiselela Umtapo Wolwazi

Ukwakhiwa komtapo wolwazi

Ukulandelana

Ukulandelana

Ukuhlaziywa kwedatha

Ukuhlaziywa kwedatha

Izinsizakalo Zangemva Kokuthengisa

Izinsizakalo zangemuva kokuthengisa

  • Okwedlule:
  • Olandelayo:

    • 流程图 12

    Ukuhlaziywa okuyinhloko kufaka phakathi:

     

    1) Ukuhlanganiswa kwe-T2T Genome

    ● I-genome ye-T2T ibhekisela ku-genome ene-“0 gaps” lapho okungenani i-chromosome eyodwa ihlanganiswa ngokuphelele kusukela ku-telomere kuya ku-telomere.

    ● Ukusebenzisa ukufundwa kwe-CCS okunembe kakhulu kanye nokufundwa okude kakhulu kwe-ONT:

    * Khiqiza i-genome ye-contig v1 nge-hybrid assembly usebenzisa i-hifiasm (v0.25.0).

    * Susa ama-plastid nama-sequence angcolile yi-BLAST ngokumelene nesizindalwazi se-NT.

    * I-scaffold ihlangana ibe yi-chromosome-scale assembly kusetshenziswa idatha ye-Hi-C ene-3D-DNA.

    * Gcwalisa ama-telomere angekho ngokuhlanganiswa kwendawo nge-ONT reads ukuze uthole i-genome yokugcina ye-T2T.

     

    2) Ukuhlolwa Kokuhlanganiswa

    ● Ukuhlolwa kwe-BUSCO

    I-BUSCO v5.2.1 (Benchmarking Universal Single-Copy Orthologs) yakha amasethi ezakhi zofuzo anekhophi eyodwa ezinhlotsheni ezinkulu zokuziphendukela kwemvelo ngokusekelwe kusizindalwazi se-OrthoDB 10. I-genome ehlanganisiwe ihlolwa ngokuqondanisa ngokumelene naleli sethi yezakhi zofuzo, ngokusekelwe kusilinganiso sokufanisa kanye nobuqotho.

    Ingxenye ephezulu ye-“Complete BUSCOs” ikhombisa ukuphelela okuphezulu kokuhlanganiswa kwe-genome.

     

    ● Ufunda Ukumapha

    Qondanisa ukufundwa okufushane okuvela ekulandeleni kwesizukulwane esilandelayo (isb., i-Illumina) ne-genome ehlanganisiwe usebenzisa i-bwa. Qondanisa ukufundwa okude kwesizukulwane sesithathu ne-genome ehlanganisiwe usebenzisa i-Minimap2.

    Ukuphelela kwe-genome ehlanganisiwe kanye nokufana kokumbozwa kokulandelana kuhlolwa ngokusekelwe esilinganisweni semephu, isilinganiso sokumbozwa kwe-genome, kanye nokusatshalaliswa kokujula.

     

    ● Ukuhlolwa kwe-Genome QC

    Hlola ukuhlanganiswa usebenzisa i-Merqury ngokuqhathanisa ukufundwa kokulandelana okunembe kakhulu kwe-k-mers kanye nokuhlanganiswa kwe-genome ukuze uthole ikhwalithi yokuvumelana (QV).

    Amanani aphezulu ekhwalithi abonisa ukunemba okuphezulu kwe-genome ehlanganisiwe.

     

    ● Ukuhlolwa kwe-Genome LAI

    I-LAI (i-LTR Assembly Index) ihlola ubuqotho bokuhlanganiswa kwe-genome njengesilinganiso sokulandelana kwe-LTR retrotransposon okungaguquki kulandelelwano lwe-LTR oluphelele. Ukulandelana kwe-LTR-RT okufanelekile kutholakala kusetshenziswa i-LTR_FINDER (v1.0.7) kanye ne-LTRharvest (v1.5.9), bese kuhlungwa futhi kuhlanganiswe kusetshenziswa i-LTR_retriever (v2.8) ukuthola ama-retrotransposon e-LTR athembekile kakhulu futhi kubalwe i-LAI.

    Ngokusho kokushicilelwa konjiniyela be-LAI, amanani e-LAI ahlukaniswe amazinga amathathu:

    Okusalungiswa (0 ≤ LAI <10), Inkomba (10 ≤ LAI <20), kanye negolide (LAI ≥ 20).

     

    ● Ukuhlonza ama-Telomere nama-Centromere

    Thola amayunithi aphindaphindayo e-telomere ku-genome usebenzisa i-TIDK. Thola ukulandelana kwe-telomere bese uthola ulwazi lwendawo usebenzisa i-FindTelomeres ngokusekelwe kuma-motif aphindaphindayo.

    Thola ukuphindaphinda okungenzeka kwe-centromeric usebenzisa i-Centromics enokufundwa okude kwesizukulwane sesithathu, bese uphinda ubheke ku-genome ukuze uthole izikhundla kanye nokulandelana kwe-centromere.

     

    1) Imephu ye-Chromosome ye-Genome

    产品主图1

    2)Izikhundla zeTelomere kuGenome

    Chr

    Ubude be-Ch(bp)

    Ukuqala_Okuphezulu(bp)

    Ukuphela_Okuphezulu(bp)

    Ubude_Obuphezulu(bp)

    Ukuqala_Okuphansi(bp)

    Ukuphela_Komfula(bp)

    Ubude_Obuphansi(bp)

    I-Chr01

    55,340,768

    53

    2,036

    1,984

    55,338,794

    55,340,768

    1,975

    Chr02

    56,588,289

    1

    2,760

    2,760

    56,584,191

    56,588,289

    4,099

    Chr03

    46,886,733

    20

    3,001

    2,982

    46,881,994

    46,886,733

    4,740

    Chr04

    49,401,798

    1

    2,143

    2,143

    49,399,160

    49,401,798

    2,639

    Chr05

    45,855,317

    10

    3,043

    3,034

    45,852,809

    45,855,317

    2,509

    Chr06

    45,285,625

    1

    3,268

    3,268

    45,283,427

    45,285,625

    2,199

    Chr07

    48,122,726

    1

    2,317

    2,317

    48,120,519

    48,122,726

    2,208

    Ni-ote:

    I-Chromosome ID

    Ubude_be-Chr (bp): Ubude be-Chromosome

    I-Upstream_Start (bp): Indawo yokuqala ye-telomere ephezulu ku-chromosome

    I-Upstream_End (bp): Indawo yokugcina ye-telomere ephezulu ku-chromosome

    Ubude_Buphezulu (bp): Ubude be-telomere ephezulu ku-chromosome

    I-Downstream_Start (bp): Indawo yokuqala ye-telomere engezansi ku-chromosome

    I-Downstream_End (bp): Indawo yokugcina ye-telomere engezansi ku-chromosome

    Ubude_Bephansi (bp): Ubude be-telomere engezansi ku-chromosome

    3)Izikhundla zeCentromere ku-Genome

    Chr

    Ubude_be-Chr(bp)

    I-Centromics_Start(bp)

    I-Centromics_End(bp)

    I-Chr01

    55,340,768

    18,943,204

    23,005,555

    Chr02

    56,588,289

    28,114,720

    30,677,916

    Chr03

    46,886,733

    24,487,558

    24,929,326

    Chr04

    49,401,798

    20,976,875

    22,563,388

    Chr05

    45,855,317

    18,578,095

    19,715,924

    Chr06

    45,285,625

    19,398,436

    19,950,173

    Chr07

    48,122,726

    26,390,720

    27,913,284

    Qaphela:

    I-Chromosome ID

    Ubude_be-Chr (bp): Ubude be-Chromosome

    I-Centromere_Start (bp): Indawo yokuqala ye-centromere ku-chromosome

    I-Centromere_End (bp): Indawo yokugcina ye-centromere ku-chromosome

    4) Izibalo Zegebe Lemiphumela Yokuhlanganiswa

    Iqembu

    Inombolo_Yesikhala

    I-Len

    I-Chr01

    0

    55,340,768

    Chr02

    0

    56,588,289

    Chr03

    0

    46,886,733

    Chr04

    0

    49,401,798

    Chr05

    0

    45,855,317

    Chr06

    0

    45,285,625

    Chr07

    0

    48,122,726

    Ingqikithi (Isilinganiso %)

    0

    347,481,256(100.00)

    Ni-ote:

    Iqembu: I-Chromosome ID

    Inombolo_Yesikhala: Inani lezikhala ku-chromosome

    I-Len (bp): Ubude be-Chromosome

    5) Ukuhlolwa kwe-Genome LAI

    i-genome.LAI

    Chr

    Ubude be-Chr(bp)

    Iqinile

    Isiyonke

    raw_LAI

    I-LAI

    i-genome_yonke

    347,481,256

    0.046

    0.36

    12.94

    15.18

    Qaphela: Ngokusho kokushicilelwa ngabathuthukisi be-LAI, amanani e-LAI ahlukaniswe ngezigaba ezintathu: Okusalungiswa (0 ≤ LAI < 10), Ireferensi (10 ≤ LAI < 20), kanye neGolide (LAI ≥ 20).

    Ubude be-Ch (bp): Ubude be-Chromosome

    Okungaphelele: Ingxenye yama-LTR-RT aphelele ku-genome

    Isiyonke: Ingxenye yama-LTR aphelele ku-genome

    raw_LAI = Okungaphelele / Isiyonke × 100

    I-LAI: Inani le-LAI elilungisiwe

    Hlola intuthuko eyenziwe yizinsizakalo zokuhlanganisa i-de novo genome ze-BMKGene ngeqoqo lezincwadi ezikhethiwe:

     

    T2T Gi-enome

    U-Liu, u-Shoucheng et al."Ukuhlanganiswa kwezakhi zofuzo ze-telomere-kuya-telomere kanye nedatha ye-multi-omic kunikeza ukuqonda ngokuvela kukakolweni wesinkwa se-hexaploid."Izakhi zofuzo zemvelo ivolumu. 57,4 (2025): 1008-1020. doi:10.1038/s41588-025-02137-x

    Yao, Xue-Feng et al."Ukuhlanganiswa okuphelele kwe-genome yerayisi yaseJaponica i-Zhonghua 11."Ukuxhumana kwezitshalo umqulu 6,10 (2025): 101463. doi:10.1016/j.xplc.2025.101463

    Lv, Zhiyuan nabanye."Ukuhlangana kwe-genome okuseduze kwe-telomere kuya ku-telomere kwe-Camellia pitardii."Idatha yesayensi ivolumu. 12,1 1422. 14 Aug. 2025, doi:10.1038/s41597-025-05764-5

    UDu, uHaiyuan nabanye."Ukuhlanganiswa kwezakhi zofuzo okucishe kuphelele kwe-Fragaria iinumae."I-BMC genomics ivolumu. 26,1 253. 14 Mar. 2025, doi:10.1186/s12864-025-11440-0

    U-Chen, u-Weikai et al."Ukuhlanganiswa okuphelele kwezakhi zofuzo zikaNicotiana benthamiana kwembula isimo sezakhi zofuzo kanye nese-epigenetic sama-centromere."Izitshalo zemvelo ivolumu. 10,12 (2024): 1928-1943. doi:10.1038/s41477-024-01849-y

     

    I-Genome ye-T2T exazululwe yi-Haplotype

    Khan, Falak Sher et al. "Ama-genome e-T2T angenamagebe axazululwe yi-Haplotype e-Cabernet Sauvignon, uhlobo lwe-winegrape."Idatha yesayensi, 10.1038/s41597-026-06910-3. 26 Feb. 2026, doi:10.1038/s41597-026-06910-3

     

    I-Genome ye-T2T + I-Genome Eqhathanisayo

    UHong, uLin nabanye. "Ukwakhiwa nokuhlaziywa kwama-genome e-telomere-kuya-telomere kwama-orange amabili amnandi: iLonghuihong neNewhall (i-Citrus sinensis)."Isayensi yeGigaumqulu 13 (2024): giae084. doi:10.1093/gigascience/giae084

    ULi, uXiao-Jie nabanye. “Ukuhlaziywa kwe-genome ye-telomere-to-telomere ye-red carrot TXH4 kuchaza indima ye-DcLCYE kanye ne-DcLCYB1 ekuqongeleleni kwe-lycopene kwi-carrot.”Ucwaningo lwezolimoivolumu. 12,11 uhaf192. 29 Jul. 2025, doi:10.1093/hr/uhaf192

     

    I-Genome ye-T2T + i-Pangenome

    UWang, uXiaojing nabanye. “I-genome ye-T2T, ukuhlaziywa kwe-pan-genome, kanye nezakhi zofuzo zokuphendula ukucindezeleka kokushisa ezinhlotsheni ze-Rhododendron.”iMetaivolumu. 4,2 e70010. 5 Mar. 2025, doi:10.1002/imt2.70010

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