Ukwehliswa Kokumelwa Kwe-Bisulfite Sequencing (RRBS)
Ukwehliswa kokumelwa kwe-bisulfite sequencing (RRBS) kuncike ekucebiseni izifunda ezicebile eziqhingini ze-CpG ngokuhlukaniswa kwe-MspI kulandelwe ukukhethwa kosayizi wezingcezu ezingama-200-500/600 bps. Ngenxa yalokho, yizifunda eziseduze neziqhingi ze-CpG kuphela ezihlelwa ngokulandelana, kuyilapho lezo ezineziqhingi ze-CpG ezikude zingafakwanga ekuhlaziyweni. Le nqubo, ehlanganiswe nokwehliswa kwe-bisulfite sequencing, ivumela ukutholakala kwe-methylation ye-DNA enesinqumo esiphezulu, kanti indlela yokwehliswa kwe-sequencing, i-PE150, igxile ngqo emaphethelweni ezi-insert esikhundleni se-middle, okwandisa ukusebenza kahle kwe-methylation profiling.
Le ndlela ivele njengendlela engcono kakhulu futhi ephumelelayo esikhundleni se-Whole Genome Bisulfite Sequencing (WGBS) ocwaningweni lwe-DNA methylation. Ngenkathi i-WGBS inikeza ukuqonda okuphelele ngokuhlola lonke i-genome ngesisombululo esisodwa, izindleko zayo eziphakeme zingaba yisici esinqunyelwe, i-RRBS eyinciphisa ngokuhlelekile le nselele ngokuhlaziya ngokukhetha ingxenye emele i-genome. Le ndlela iyithuluzi elibalulekile elivumela ucwaningo lwe-DNA methylation olungabizi futhi luthuthukise ulwazi lwezindlela ze-epigenetic.
Izici Zesevisi
● Ukulandelana kwezithombe ku-Illumina NovaSeq.
● Kudinga i-genome ebhekisela kuyo.
● I-Lambda DNA isetshenziselwa ukuqapha ukusebenza kahle kokuguqulwa kwe-bisulfite.
● Ukusebenza kahle kokugaya kwe-MspI nakho kuyaqashelwa.
● Ukugaya ama-enzyme kabili kumasampula ezitshalo.
Izinzuzo Zesevisi
●Indlela Engabizi Kakhulu Nephumelelayo Esikhundleni se-WGBS: Kwenza ukuthi ukuhlaziywa kwenziwe ngezindleko eziphansi futhi kube nezidingo zesampula eziphansi.
●Ipulatifomu Ephelele:Sinikeza isevisi enhle kakhulu kusukela ekucubungulweni kwamasampula, ukwakhiwa komtapo wolwazi, kanye nokulandelana kwawo kuya ekuhlaziyweni kwe-bioinformatics.
●Ubuchwepheshe Obubanzi: Siphumelele ezinhlobonhlobo zezinhlobo. I-BMKGENE iletha ulwazi olungaphezu kweminyaka eyishumi, ithimba lokuhlaziya elinekhono eliphezulu, okuqukethwe okuphelele, kanye nokusekelwa okuhle kakhulu ngemuva kokuthengisa.
Imininingwane Yesevisi
| Umtapo Wolwazi | Isu Lokulandelana | Ukukhishwa kwedatha okunconyiwe | Ikhwalithi yokulawula |
| Umtapo wolwazi ohlungiwe yi-MspI kanye ne-Bisulfite | I-Illumina PE150 | 8 Gb | Q30 ≥ 85% Ukuguqulwa kwe-Bisulfite > 99% Ukusebenza kahle kokusika kwe-MspI > 95% |
Izidingo Zesampula
| Ukugxila (ng/µL) | Inani eliphelele (µg) |
| |
| I-DNA ye-Genomic | ≥ 30 | ≥ 1 | Ukonakala noma ukungcola okulinganiselwe |
Ukugeleza Komsebenzi Wesevisi
Ukulethwa kwesampula
Ukwakhiwa komtapo wolwazi
Ukulandelana
Ukuhlaziywa kwedatha
Izinsizakalo zangemuva kokuthengisa
Ukuhlaziywa kwe-BI kuhlanganisa:
● Ukulawulwa kwekhwalithi yokulandelana okungahluziwe;
● Ukuhlela i-genome ebhekisele kuyo;
● Ukutholwa kwezisekelo ze-methylated ezingama-5mC kanye nokuhlonza i-motif;
● Ukuhlaziywa kokusatshalaliswa kwe-methylation kanye nokuqhathaniswa kwesampula;
● Ukuhlaziywa Kwezindawo Ezine-Methylated Ehlukile (ama-DMR);
● Isichasiselo esisebenzayo sezakhi zofuzo ezihlotshaniswa nama-DMR.
Ukulawulwa kwekhwalithi: ukusebenza kahle kokugaya ukudla (ekuhlolweni kwe-genome)
Ukulawulwa kwekhwalithi: ukuguqulwa kwe-bisulfite (ekukhishweni kolwazi lwe-methylation)
Imephu ye-Methylation: Ukusatshalaliswa kwe-genome ye-methylation ye-5mC
Ukuqhathanisa isampula: Ukuhlaziywa Kwezingxenye Eziyinhloko
Ukuhlaziywa Kwezifunda Ezine-Methylated Ngokuhlukile (DMRs): imephu yokushisa
Hlola intuthuko yocwaningo eyenziwe yizinsizakalo zokulandelana kwe-genome bisulfite ze-BMKGene ngokusebenzisa iqoqo lezincwadi ezikhethiwe.
ULi, Z. et al. (2022) 'Ukuhlelwa kabusha kokwethembeka okuphezulu kumaseli afana neLeydig ngokusebenzisa ukusebenza kwe-CRISPR kanye nezinto ze-paracrine',I-PNAS Nexus, 1(4). doi: 10.1093/PNASNEXUS/PGAC179.
UTian, H. et al. (2023) 'Ukuhlaziywa kwe-genome-wide DNA methylation yokwakheka komzimba kumawele aseShayina angama-monozygotic',Ijenali YaseYurophu Yophenyo Lwezokwelapha, 53(11), p. e14055. doi: 10.1111/ECI.14055.
UWu, Y. et al. (2022) 'I-DNA methylation kanye nesilinganiso sesinqe nesinqe: ucwaningo lokuhlobana kwe-epigenome kumawele aseShayina angama-monozygotic',Ijenali Yophenyo Lwe-Endocrinological, 45(12), amakhasi 2365-2376. doi: 10.1007/S40618-022-01878-4.
