Whakaitihia te Whakaaturanga Bisulfite Sequencing (RRBS)
Kua puta ake te Whakahekenga Whakaitinga Bisulfite Sequencing (RRBS) hei utu utu me te whai hua ki te Whole Genome Bisulfite Sequencing (WGBS) i roto i te rangahau methylation DNA. Ahakoa e whakarato ana a WGBS i nga tirohanga matawhānui ma te tirotiro i te ira ira katoa i te taumira turanga kotahi, ko te utu nui ka waiho hei take whakaiti. Ka taea e RRBS te whakaiti i tenei wero ma te whiriwhiri i tetahi waahanga o te genome. Ko tenei tikanga e whakawhirinaki ana ki te whakarangatira i nga rohe-motu nui o CpG na roto i te wehenga MspI whai muri i te kowhiringa rahi o nga kongakonga 200-500/600 bps. No reira, ko nga rohe anake e tata ana ki nga moutere CpG ka raupapahia, engari ko nga rohe kei tawhiti atu i nga moutere CpG kaore i uru ki te tātari. Ko tenei tukanga, me te raupapa bisulfite, ka taea te kimi teitei o te methylation DNA, me te huarahi raupapa, PE150, e arotahi ana ki nga pito o nga whakauru kaore i te waenganui, me te whakanui ake i te pai o te methylation profiling. Ko te RRBS he taputapu utu nui e taea ai te rangahau methylation DNA whai hua me te ahu whakamua i te matauranga mo nga tikanga epigenetic.
Nga waahanga ratonga
● Me whai ira tohutoro.
● Ka whakamahia te DNA Lambda ki te aro turuki i te pai o te hurihanga bisulfite.
● Ka aro turukihia te pai o te nakunaku MspI.
● Te nakunaku whākōkī rua mō ngā tauira tipu.
● Whakaraupapa i runga i te Illumina NovaSeq.
Painga Ratonga
●Utu-whai hua me te whai hua ki te WGBS: kia taea ai te tātaritanga te whakahaere i te utu iti me te iti o nga whakaritenga tauira.
●Paerewa Whakaoti:whakarato ratonga pai kotahi-mutu mai i te tukatuka tauira, te hanga whare pukapuka, me te raupapatanga ki te tātari bioinformatics.
●Tohunga Nui: me nga kaupapa raupapa RRBS kua oti pai puta noa i nga momo momo kanorau, ka kawea mai e BMKGENE mo te tekau tau o te wheako, he roopu tātari tino mohio, he korero matawhānui, me te tautoko pai i muri i te hokonga.
Whakatakotoranga Ratonga
| Whare Pukapuka | Rautaki Raupapa | Putanga raraunga e taunaki ana | Mana kounga |
| MspI kua keri me te wharepukapuka rongoa Bisulfite | Ilumina PE150 | 8 Gb | Q30 ≥ 85% Te hurihanga bisulfite > 99% MspI pai tapahi > 95% |
Nga Whakaritenga Tauira
| Kukū (ng/µL) | Tapeke tapeke (µg) |
| |
| DNA genomic | ≥ 30 | ≥ 1 | He iti te whakaheke, te poke ranei |
Te Rerenga Mahi Ratonga
Te tuku tauira
Te hanga whare pukapuka
Whakaraupapa
Te tātari raraunga
Nga ratonga muri-hoko
Kei roto ko te tātaritanga e whai ake nei:
● Raw sequencing mana kounga;
● Mahere ki te tohutoro iraira;
● Te kitenga o nga turanga methylated 5mC me te tohu tohu;
● Te tātaritanga o te tohatoha methylation me te whakataurite tauira;
● Tātaritanga o ngā Rohe Methylated Differentially (DMRs);
● Tuhituhi mahi o nga ira e hono ana ki nga DMR.
Te mana o te kounga: te mahi nakunaku (i roto i te mapi ira ira)
Mana kounga: te hurihanga bisulfite (i te tangohanga korero methylation)
Mapi methylation: 5mC methylation tohatoha ira-whanui
Tauira whakataurite: Te Tirohanga Waa Matua
Te tātaritanga o nga Rohe Methylated Differentially Methylated (DMRs): mahere wera
Torotoro i nga mahi rangahau i whakahaerea e nga ratonga whakaraupapa bisulfite genome katoa a BMKGene na roto i te kohinga whakaputanga.
Li, Z. et al. (2022) 'Whakahōtaka hou ki roto i nga pūtau rite-Leydig na te whakahohenga CRISPR me nga mea paracrine',PNAS nohonga, 1(4). doi: 10.1093/PNASNEXUS/PGAC179.
Tian, H. et al. (2023) 'Te tātari methylation DNA-Genome-wide mo te hanganga tinana i roto i nga mahanga monozygotic Hainamana',Te Tuhituhi a te Pakeha mo te Tirohanga Haumanu, 53(11), wh. e14055. doi: 10.1111/ECI.14055.
Wu, Y. et al. (2022) 'DNA methylation me te waist-to-hip ratio: he rangahau hononga epigenome-wide i roto i nga mahanga monozygotic Hainamana',Journal of Endocrinological Investigation, 45(12), api 2365–2376. doi: 10.1007/S40618-022-01878-4.
