I-Specific-Locus Amplified Fragment Sequencing (SLAF-Seq)
Ukuhamba komsebenzi
Uhlelo Lokusebenza
Izici Zesevisi
● Ukulandelanisa ku-NovaSeq nge-PE150.
● Ukulungiswa komtapo wezincwadi onamabhakhodi amabili, okuvumela ukuhlanganisa amasampula angaphezu kuka-1000.
● Le nqubo ingasetshenziswa noma ngaphandle kwe-genome yereferensi, enamapayipi ahlukene we-bioinformatic esimweni ngasinye:
Nge-genome yereferensi: ukutholwa kwe-SNP kanye ne-InDel
Ngaphandle kwegenome yereferensi: ukuhlanganisa isampula nokutholwa kwe-SNP
● Kuku-siliconeInhlanganisela ye-enzyme ekhawulelwe ngaphambili yesigaba sokuklama iyahlolwa ukuze kutholwe lezo ezikhiqiza ukusatshalaliswa okufanayo kwamathegi e-SLAF ku-genome.
● Phakathi nokuhlolwa kwangaphambilini, inhlanganisela yama-enzyme amathathu ayahlolwa kumasampuli angu-3 ukuze kukhiqizwe imitapo yolwazi ye-SLAF engu-9, futhi lolu lwazi lusetshenziselwa ukukhetha inhlanganisela ye-enzyme ekhawulelwe efanele yephrojekthi.
Izinzuzo Zesevisi
●Ukutholwa Komaka Okuphakeme Wofuzo: Ukuhlanganisa uhlelo oluphezulu lwebhakhodi ephindwe kabili kuvumela ukulandelana ngesikhathi esisodwa kwesibalo sabantu abaningi, futhi ukukhuliswa okuqondene nendawo ethile kuthuthukisa ukusebenza kahle, kuqinisekisa ukuthi izinombolo zomaka zihlangabezana nezidingo ezihlukahlukene zemibuzo yocwaningo eyahlukahlukene.
● Ukuncika Okuphansi ku-Genome: Ingasetshenziswa ezinhlotsheni ezine-genome eyisithenjwa noma ezingenayo.
●I-Flexible Scheme Design: I-enzyme eyodwa, i-dual-enzyme, ukugaya kwama-enzyme amaningi, nezinhlobo ezihlukahlukene zama-enzyme zingakhethwa ukuze kuhlangatshezwane nezinhloso zocwaningo noma izinhlobo ezihlukahlukene. Iku-siliconeukwakheka kwangaphambilini kwenziwa ukuze kuqinisekiswe ukwakheka okuhle kwe-enzyme.
● Ukusebenza Okuphezulu Kwe-Enzymatic Digestion: Ukwenziwa kwe-ku-siliconeukuklama kwangaphambilini kanye nomklamo wangaphambili oqinisekisiwe ngokusatshalaliswa okulinganayo komaka be-SLAF ku-chromosome (ithegi engu-1 SLAF/4Kb) nokunciphisa ukulandelana okuphindaphindayo (<5%).
●Ubungcweti Olubanzi: Ithimba lethu liletha ingcebo yesipiliyoni kuyo yonke iphrojekthi, enomlando wokuvala amaphrojekthi we-SLAF-Seq angaphezu kuka-5000 kumakhulu ezinhlobo zezinto eziphilayo, okuhlanganisa izitshalo, izilwane ezincelisayo, izinyoni, izinambuzane, nezinto eziphilayo zasemanzini.
● Ukuhamba komsebenzi okuzithuthukisayo kwe-Bioinformatic: I-BMKGENE ithuthukise ukuhamba komsebenzi okuhlanganisiwe kwe-bioinformatic ye-SLAF-Seq ukuze kuqinisekiswe ukwethembeka nokunemba kokuphumayo kokugcina.
Imininingwane Yesevisi
| Uhlobo lokuhlaziya | Isilinganiso sabantu esinconyiwe | Isu lokulandelanisa | |
| Ukujula kokulandelana komaka | Inombolo yethegi | ||
| I-Genetic Maps | Abazali abangu-2 kanye nezingane ezingaphezu kuka-150 | Abazali: 20x WGS Ukukhipha: 10x | Usayizi weGenome: <400 Mb: I-WGS iyanconywa <1Gb: 100K omaka 1-2Gb:: 200K omaka > 2Gb: 300K omaka Amathegi aphezulu angama-500k |
| Izifundo ze-Genome-Wide Association (GWAS) | ≥200 amasampula | 10x | |
| I-Genetic Evolution | ≥30 amasampuli, namasampuli >10 avela eqenjini ngalinye elincane | 10x | |
Izidingo Zesevisi
Ukugxila ≥ 5 ng/µL
Inani eliphelele ≥ 80 ng
I-Nanodrop OD260/280=1.6-2.5
Ijeli ye-Agarose: akukho noma ukuwohloka okulinganiselwe noma ukungcoliswa
Ukulethwa Kwesampula Okunconyiwe
Isitsha: 2 ml centrifuge tube
(Kumasampuli amaningi, sincoma ukuthi ungayigcini ku-ethanol)
Isampula yokulebula: Amasampuli adinga ukulebula ngokucacile futhi afane nefomu lesampula lolwazi elithunyelwe.
Ukuthunyelwa: Iqhwa elomile: Amasampula adinga ukupakishwa ezikhwameni kuqala futhi agqitshwe eqhweni elomile.
Uhlelo lokusebenza lwesevisi
Isampula ye-QC
Ukuhlolwa komshayeli
SLAF-ukuhlola
Ukulungiselela Umtapowolwazi
Ukulandelanisa
Ukuhlaziywa Kwedatha
Amasevisi angemuva kokuthengisa
Kufaka ukuhlaziya okulandelayo:
- Ilandelanisa idatha ye-QC
- Ukuthuthukiswa kwethegi ye-SLAF
Imephu yokubhekisela ku-genome
Ngaphandle kwe-genome eyireferensi: ukuhlanganisa
- Ukuhlaziywa kwamathegi e-SLAF.: izibalo, ukusatshalaliswa ku-genome yonkana
- Ukutholwa komaka: SNP, InDel, SNV, CV call and annotation
Ukusatshalaliswa kwamathegi e-SLAF kuma-chromosome:
Ukusatshalaliswa kwama-SNP kuma-chromosome:
Isichasiselo se-SNP
| Unyaka | Ijenali | IF | Isihloko | Izinhlelo zokusebenza |
| 2022 | Ukuxhumana kwemvelo | 17.694 | Isisekelo se-genomic se-giga-chromosomes kanye ne-giga-genome ye-peony yesihlahla I-Paeonia ostii | I-SLAF-GWAS |
| 2015 | I-Phytologist entsha | 7.433 | Izinyathelo zasekhaya ziqinisa izifunda ze-genomic ezibalulekile ku-agronomic ubhontshisi wesoya | I-SLAF-GWAS |
| 2022 | Ijenali Yocwaningo Oluthuthukile | 12.822 | I-Genome-wide-wide introgressions yokwenziwa kwe-Gossypium barbadense ku-G. hirsutum veza i-loci ephezulu yokuthuthukisa kanyekanye ikhwalithi yefayibha kakotini kanye nesivuno izici | I-SLAF-Evolutionary genetics |
| 2019 | Isitshalo samangqamuzana | 10.81 | I-Population Genomic Analysis kanye ne-De Novo Assembly Yembula Umsuka Weedy Ilayisi njengomdlalo wokuziphendukela kwemvelo | I-SLAF-Evolutionary genetics |
| 2019 | I-Nature Genetics | 31.616 | Ukulandelana kwe-genome kanye nokuhlukahluka kofuzo kwe-carp evamile, i-Cyprinus carpio | Imephu ye-SLAF-Linkage |
| 2014 | I-Nature Genetics | 25.455 | I-genome ye-peanut etshaliwe inikeza ukuqonda nge-karyotypes ye-legume, i-polyploid ukuziphendukela kwemvelo kanye nokufuywa kwezitshalo. | Imephu ye-SLAF-Linkage |
| 2022 | Plant Biotechnology Journal | 9.803 | Ukukhonjwa kwe-ST1 kuveza ukukhetha okubandakanya ukugitsheliswa kwembewu yokuma kwembewu kanye nokuqukethwe kwamafutha ngesikhathi sokwenziwa kwe-soya | Ukuthuthukiswa kwe-SLAF-Marker |
| 2022 | Ijenali Yamazwe Ngamazwe Yesayensi Yamangqamuzana | 6.208 | Ukuhlonzwa kanye Nokuthuthukiswa Kwe-DNA Marker ye-Wheat-Leymus mollis 2Ns (2D) I-Disomic Chromosome Substitution | Ukuthuthukiswa kwe-SLAF-Marker |
| Unyaka | Ijenali | IF | Isihloko | Izinhlelo zokusebenza |
| 2023 | Imingcele kusayensi yezitshalo | 6.735 | Imephu ye-QTL kanye nokuhlaziywa kwe-transcriptome kokuqukethwe ushukela ngesikhathi sokuvuthwa kwezithelo zePyrus pyrifolia | Imephu Yofuzo |
| 2022 | Plant Biotechnology Journal | 8.154 | Ukukhonjwa kwe-ST1 kuveza ukukhetha okubandakanya ukugitsheliswa kwembewu ye-morphology kanye nokuqukethwe kukawoyela ngesikhathi sokukhiqizwa kukabhontshisi wesoya.
| Ukushaya kwe-SNP |
| 2022 | Imingcele kusayensi yezitshalo | 6.623 | I-Genome-Wide Association Mapping of Hulless Barely Phenotypes Emvelweni Yesomiso.
| I-GWAS |
