Kutevedzana kweZvikamu Zvakatsanangurwa zveLocus (SLAF-Seq)
Kufamba kwebasa
Basa iri rine dhizaini yakagadzirwa kare mu silico kuti rive nechokwadi chekuti ma enzymes akasarudzwa zvakanaka pakugadzirira raibhurari.
Chirongwa cheUnyanzvi
Zvimiro zveBasa
● Kutevedzana kweNovaSeq nePE150.
● Kugadzirira raibhurari nekushandisa mabarcode maviri, zvichibvumira kuunganidzwa kwemasampuli anopfuura chiuru.
● Zvisingabatanidzwi ne genome reference:
Nekutsanangurwa kwe genome: kuwanikwa kweSNP neInDel
Pasina reference genome: sample clustering uye kuwanikwa kweSNP
● Mukatimu-silicoPadanho rekutanga rekugadzira, musanganiswa wemaenzyme akawanda ekudzivirira anoongororwa kuti awane ayo anogadzira kupararira kwakafanana kwematagi eSLAF mugenome.
● Munguva yekuedza kusati kwaitwa, musanganiswa wema enzyme matatu unoedzwa musampuli nhatu kuti pagadzirwe maraibhurari mapfumbamwe eSLAF, uye ruzivo urwu runoshandiswa kusarudza musanganiswa wema enzyme akakodzera wepurojekiti.
Zvakanakira Basa
●Kuwanikwa kweHigh Genetic Marker: Tinobatanidza sisitimu ine mabarcode maviri ane simba repamusoro inobvumira kurongeka kwevanhu vakawanda panguva imwe chete, uye kuwedzera kwenzvimbo kwakanangana nekuwedzera kushanda zvakanaka, tichiona kuti ma tag numbers anosangana nezvinodiwa zvakasiyana-siyana zvemibvunzo yakasiyana-siyana yekutsvagisa.
● Kusavimba neGenome: Inogona kushandiswa kumarudzi ane kana asina genome reference.
●Dhizaini Yechirongwa Chinochinjika: Kugayiwa kwemaenzyme rimwe chete, maenzyme maviri, kugaya kwemaenzyme akawanda, uye mhando dzakasiyana dzemaenzyme zvinogona kusarudzwa kuti zvienderane nezvinangwa zvakasiyana zvekutsvaga kana mhando.
● Kushanda Kwakanyanya Mukugaya KwemaEnzymatic: Kutungamirwa kwemu-silicoKugadzirwa kwekutanga uye kuyedzwa kwekutanga kunovimbisa dhizaini yakanaka nekupararira kwakaenzana kwematagi eSLAF pachromosome (1 SLAF tag/4Kb) uye kuderedzwa kwekutevedzana kunodzokororwa (<5%).
●Unyanzvi Hwakanyanya: Tinounza ruzivo rwakakura kupurojekiti yega yega, nenhoroondo yekupedzisa mapurojekiti eSLAF-Seq anopfuura 5000 pamarudzi ezvisikwa, kusanganisira zvirimwa, mhuka dzinoyamwisa, shiri, zvipembenene, uye zvipenyu zvemumvura.
● Kufamba Kwebasa reBioinformatic rakazvigadzira: Takagadzira nzira yakabatana yekushanda kweSLAF-Seq kuti tive nechokwadi chekuvimbika uye kururama kwemhedzisiro yekupedzisira.
Tsanangudzo dzeBasa
| Rudzi rwekuongorora | Chiyero chehuwandu hwevanhu chinokurudzirwa | Maitiro ekuteverana | |
| Kudzika kwekutevedzana kwema tag | Nhamba yeTag | ||
| Mepu dzeMajini | Vabereki vaviri nevana vanopfuura zana nemakumi mashanu | Vabereki: 20x WGS Kubvisa: 10x | Saizi yemajini: <400 Mb: WGS inokurudzirwa <1Gb: 100K tags 1-2Gb:: 200K tags >2Gb: 300K tags Ma tag epamusoro e500k |
| Zvidzidzo zveGenome-Wide Association (GWAS) | ≥200 sampuro | 10x | |
| Kushanduka Kwemajini | ≥30 sampuro, ne >10 sampuro kubva kuboka rega rega | 10x | |
Zvinodiwa Pabasa
Kuwanda kwemvura ≥ 5 ng/µL
Huwandu hwese ≥ 80 ng
Nanodrop OD260/280=1.6-2.5
Gel yeAgarose: haina kana kuti haina kusvibiswa kana kusvibiswa kushoma
Muenzaniso Unokurudzirwa Kutumirwa
Chirongo: 2 ml chubhu ye centrifuge
(Kune akawanda emuenzaniso, tinokurudzira kuti usachengetedze mu ethanol)
Kunyora sampuli: Mienzaniso inofanira kunge yakanyorwa zvakajeka uye yakafanana nefomu reruzivo rwemuenzaniso rakatumirwa.
Kutumirwa: Aizi yakaoma: Mienzaniso inofanira kuiswa mumabhegi kutanga yovigwa muaizi yakaoma.
Mafambiro Ebasa
Muenzaniso weQC
Kuedza kwemutyairi wendege
Kuedza kweSLAF
Kugadzirira Raibhurari
Kutevedzana kwezvikamu
Kuongorora Data
Masevhisi eKutengesa Mushure mekutengesa
Ongororo yedu ye bioinformatics inosanganisira:
Data QC uye kuchekerera data kubvisa N-rich reads, adapter reads kana low quality reads.
Kudzora kwechipiri kunaka kwemavara akachena ekutarisa kugoverwa kwehwaro, mhando yekutevera uye kuongororwa kwedata, asiwo kutarisa kushanda zvakanaka kwekugaya uye ma inserts akawanikwa.
Kana kuverenga kwacho kwaongororwa, pane sarudzo mbiri:
- Kugadzira mepu yekutarisa genome
- Pasina genome reference: clustering
Mushure mazvo, kuongororwa kwematagi eSLAF kunoshandiswa kuita mamwe mafonero akasiyana-siyana kubatsira pakuwana zviratidzo: SNP, InDel, SNV, CV calling uye annotation
Kugoverwa kwematagi eSLAF pamachromosomes:
Kupararira kweSNPs pama chromosomes:
Chinyorwa cheSNP
Jiang S, Li S, Luo J, Wang X naShi C (2023) Kuongororwa kweQTL uye kuongororwa kwehuwandu hweshuga panguva yekuibva kwemicheroPyrus pyrifolia.Mberi. Sainzi yezvirimwa.14:1137104. doi: 10.3389/fpls.2023.1137104
Li, J., Zhang, Y., Ma, R., Huang, W., Hou, J., Fang, C., & Sun, L. (2022). Kuzivikanwa kwe st1 kunoratidza sarudzo inosanganisira kufamba netsoka kwemhando yembeu uye huwandu hwemafuta panguva yekudyara soya.Chinyorwa cheBiotechnology yezvirimwa, 20(6), 1110-1121. https://doi.org/10.1111/pbi.13791
Xu, P., Zhang, X., Wang, X.nevamwe.Kutevedzana kwemajini uye kusiyana kwemajini emukoko wekapepuru,Cyprinus carpio.Nat Genet 46, 1212–1219 (2014). https://doi.org/10.1038/ng.3098
Zhuang, W., Chen, H., Yang, M.nevamwe.Majini enzungu dzinorimwa anopa ruzivo rwakadzama nezvemhando dzenzungu, kushanduka kwepolyploid uye kuchengetwa kwezvirimwa.Nat Genet 51, 865–876 (2019). https://doi.org/10.1038/s41588-019-0402-2
| Gore | Chinyorwa | IF | Musoro | Mashandisirwo |
| 2022 | Kutaurirana kwezvisikwa | 17.694 | Hwaro hwe genomic hwe giga-chromosomes uye giga-genome yemuti peony Paeonia ostii | SLAF-GWAS |
| 2015 | Nyanzvi Itsva yePhytologist | 7.433 | Zviratidzo zvekugara mudzimba zvinosimbisa nzvimbo dzemajini dzakakosha pakurima soyabhinzi | SLAF-GWAS |
| 2022 | Chinyorwa Chekutsvagisa Kwepamusoro | 12.822 | Kupinda kweGossypium barbadense muG. hirsutum kuburikidza ne genome yese. inoratidza nzvimbo yepamusoro yekuvandudza kunaka kwekotoni uye goho panguva imwe chete hunhu | SLAF-Majini ekushanduka-shanduka |
| 2019 | Chirimwa cheMolecular | 10.81 | Kuongorora Magenomic eVanhu neDe Novo Assembly Zvinoratidza Kwakatangira Weedy Mupunga seMutambo weKushanduka-shanduka | SLAF-Majini ekushanduka-shanduka |
| 2019 | Majini Ezvisikwa | 31.616 | Kutevedzana kwemajini uye kusiyana kwemajini emukoko wekapfeki wakajairika, Cyprinus carpio | SLAF-Linkage mepu |
| 2014 | Majini Ezvisikwa | 25.455 | Majini enzungu dzinorimwa anopa ruzivo rwakadzama nezvemhando dzenzungu, polyploid shanduko uye kupfuya zvirimwa. | SLAF-Linkage mepu |
| 2022 | Chinyorwa cheBiotechnology yezvirimwa | 9.803 | Kuzivikanwa kweST1 kunoratidza kusarudzwa kunosanganisira kufamba netsoka dzembeu uye huwandu hwemafuta panguva yekurimwa soya | Kugadzirwa kweSLAF-Marker |
| 2022 | Chinyorwa Chepasi Pose cheSainzi dzeMolecular | 6.208 | Kuzivikanwa uye Kugadzirwa kweDNA Marker yeGorosi-Leymus mollis 2Ns (2D) Kutsiva Disomic Chromosome | Kugadzirwa kweSLAF-Marker |
| Gore | Chinyorwa | IF | Musoro | Mashandisirwo |
| 2023 | Miganhu mune sainzi yemiti | 6.735 | Kuongororwa kweQTL mepu uye transcriptome yehuwandu hweshuga panguva yekuibva kwemichero yePyrus pyrifolia | Mepu yeMajini |
| 2022 | Chinyorwa cheBiotechnology yezvirimwa | 8.154 | Kuonekwa kweST1 kunoratidza kusarudzwa kunosanganisira kufambisa bhiza remhando yembeu uye huwandu hwemafuta panguva yekudyara soya.
| Kufona kweSNP |
| 2022 | Miganhu mune sainzi yemiti | 6.623 | Kuongororwa kweGenome-Wide kweHullless Barely Phenotypes muNzvimbo Inonaya Mvura.
| GWAS |
