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I-Hi-C esekelwe ku-Chromatin Interaction
I-Hi-C iyindlela edizayinelwe ukuthwebula ukucushwa kwe-genomic ngokuhlanganisa ukuhlola okususelwe ekubeni seduze kanye nokulandelana komphumela ophezulu. Indlela isekelwe ekuxhumaneni kwe-chromatin ne-formaldehyde, okulandelwa ukugayeka kokudla kanye nokuhlanganisa kabusha ngendlela yokuthi izingcezu kuphela ezixhunywe ngokuhlanganyela zizokwakha imikhiqizo yokuhlanganisa. Ngokulandelanisa le mikhiqizo ye-ligation, kungenzeka ukufunda inhlangano ye-3D ye-genome. I-Hi-C inika amandla ukufunda ngokusatshalaliswa kwezingxenye zegenome ezipakishwe kancane (Amakhompathimenti A, i-euchromatin) futhi okungenzeka zisebenze ngokubhalwa phansi, nezifunda ezipakishwe kakhulu (amakhompathimenti B, i-Heterochromatin). I-Hi-C ingase futhi isetshenziselwe ukukhomba izizinda ezihlotshaniswa ne-Topologically Associated Domains (TADs), izifunda zofuzo ezinezakhiwo ezigoqiwe futhi okungenzeka zibe namaphethini wokukhuluma afanayo, kanye nokuhlonza amaluphu e-chromatin, izifunda ze-DNA ezihlanganiswe ndawonye ngamaphrotheni futhi evame ukucetshiswa ezintweni zokulawula. Insizakalo yokulandelana ye-Hi-C ye-BMKGene inika abacwaningi amandla okuhlola ubukhulu bendawo be-genomics, bavule izindlela ezintsha zokuqonda ukulawulwa kofuzo kanye nomthelela wako kwezempilo nezifo.
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I-Chromatin Immunoprecipitation Sequencing (ChIP-seq)
I-Chromatin Immunoprecipitation (CHIP) iyindlela esebenzisa amasosha omzimba ukuze inothise ngokukhetha amaprotheni abopha i-DNA kanye nemigomo yawo ehambisanayo ye-genomics. Ukuhlanganiswa kwayo ne-NGS kuvumela ukuhlonza okubanzi kwe-genome kokuhlosiwe kwe-DNA okuhlotshaniswa nokuguqulwa kwe-histone, izici zokuloba, namanye amaprotheni abopha i-DNA. Le ndlela eguquguqukayo inika amandla ukuqhathaniswa kwezindawo ezibophayo kuzo zonke izinhlobo zamaseli ahlukahlukene, izicubu, noma izimo. Izinhlelo zokusebenza ze-ChIP-Seq zisukela ekutadisheni imithetho yokubhala kanye nezindlela zokuthuthuka ziye ekucaciseni izindlela zezifo, okuyenza ibe ithuluzi elibalulekile lokuqonda ukubusa kokwakheka kwezwe kanye nokuthuthukiswa kwemininingwane yokwelapha.
Ipulatifomu: Illumina NovaSeq
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I-Whole genome bisulfite sequencing(WGBS)
I-Whole Genome Bisulfite Sequencing (WGBS) ime njengendlela yegolide esezingeni eliphezulu yokuhlola ngokujulile i-DNA methylation, ikakhulukazi indawo yesihlanu ku-cytosine (5-mC), umlawuli obalulekile wokuvezwa kofuzo kanye nomsebenzi wamaselula. Umgomo oyisisekelo we-WGBS uhilela ukwelashwa kwe-bisulfite, okwenza ukuguqulwa kwama-cytosine angenayo i-methylated abe yi-uracil (C kuya ku-U), kuyilapho kushiya ama-cytosine e-methylated engashintshiwe. Le nqubo inikezela ngesixazululo esisisekelo esisodwa, okuvumela abacwaningi ukuthi baphenye kabanzi nge-methylome futhi bathole amaphethini e-methylation angajwayelekile ahlobene nezimo ezahlukahlukene, ikakhulukazi umdlavuza. Ngokusebenzisa i-WGBS, ososayensi bangathola ukuqonda okungenakuqhathaniswa ku-genome-wide methylation landscapes, banikeze ukuqonda okufingqiwe kwezinqubo ze-epigenetic ezingaphansi kwezinqubo nezifo ezihlukahlukene zebhayoloji.
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I-Assay ye-Chromatin Efinyelelekayo Ye-Transposase Enokulandelanisa Okuphezulu Kokudlula (i-ATAC-seq)
I-ATAC-seq iyindlela yokulandelanisa ephezulu esetshenziselwa ukuhlaziywa kokufinyeleleka kwe-chromatin yofuzo olubanzi. Ukusetshenziswa kunikeza ukuqonda okujulile kwezinqubo eziyinkimbinkimbi zokulawula i-epigenetic emhlabeni jikelele phezu kokuchazwa kwezakhi zofuzo. Indlela isebenzisa i-hyperactive Tn5 transposase ukuze ngesikhathi esisodwa ihlukanise futhi ithege izifunda ze-chromatin ezivulekile ngokufaka ama-adaptha alandelanayo. Ukukhulisa i-PCR okulandelayo kuphumela ekudalweni kwelabhulali elandelanayo, evumela ukuhlonza okuphelele kwezifunda ze-chromatin evulekile ngaphansi kwezimo ezithile zesikhathi sesikhala. I-ATAC-seq inikeza umbono ophelele wezindawo ze-chromatin ezifinyelelekayo, ngokungafani nezindlela ezigxila kuphela kumasayithi abophezela isici sokuloba noma izifunda ezithile ezilungiswe i-histone. Ngokulandelanisa lezi zifunda ze-chromatin evulekile, i-ATAC-seq iveza izifunda okungenzeka kakhulu ukuthi zilandele ukulandelana kokulawula okusebenzayo kanye namasayithi angaba khona abopha isici sokuloba, enikeza imininingwane ebalulekile ekuguquguqukeni okuguquguqukayo kwesisho sofuzo kuyo yonke igenome.
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I-Reduced Representation Bisulfite Sequencing (RRBS)
I-Reduced Representation Bisulfite Sequencing (RRBS) ivele njengendlela engabizi futhi ephumelelayo ye-Whole Genome Bisulfite Sequencing (WGBS) ocwaningweni lwe-DNA methylation. Ngenkathi i-WGBS ihlinzeka ngemininingwane ebanzi ngokuhlola yonke i-genome ekuxazululeni kwesisekelo esisodwa, izindleko zayo eziphezulu zingaba yisici esikhawulelayo. I-RRBS inciphisa ngokuhlelekile le nselele ngokukhetha ingxenye emele yofuzo. Le ndlela yokwenza incike ekuthuthukisweni kwezifunda ezicebile eziqhingini ze-CpG nge-MspI cleavage elandelwa ukukhethwa kosayizi wezingcezu ze-bps ezingu-200-500/600. Ngokulandelayo, yizifunda kuphela eziseduze neziqhingi ze-CpG ezilandelanayo, kuyilapho lezo ezineziqhingi ezikude ze-CpG azifakwanga ekuhlaziyweni. Le nqubo, ehlangene nokulandelana kwe-bisulfite, ivumela ukutholakala kwe-high-resolution ye-DNA methylation, kanye nendlela yokulandelana, i-PE150, igxile ikakhulukazi emaphethelweni okufaka esikhundleni esiphakathi, okwandisa ukusebenza kahle kwephrofayili ye-methylation. I-RRBS iyithuluzi eliyigugu elivumela ucwaningo lwe-DNA methylation olungabizi futhi lithuthukise ulwazi lwezinqubo ze-epigenetic.
