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Hoʻopili Chromatin ma muli o Hi-C
ʻO Hi-C kahi ala i hoʻolālā ʻia e hopu i ka hoʻonohonoho genomic ma o ka hoʻohui ʻana i nā pilina pili pili kokoke a me ke kaʻina hana kiʻekiʻe. Hoʻokumu ʻia ke ʻano ma luna o ka chromatin crosslinking me formaldehyde, a ukali ʻia e ka ʻai ʻana a me ka re-ligation ma ke ʻano o nā ʻāpana i hoʻopili covalently e hana i nā huahana ligation. Ma ka hoʻonohonoho ʻana i kēia mau huahana ligation, hiki ke aʻo i ka hui 3D o ka genome. Hiki iā Hi-C ke aʻo i ka māhele ʻana o nā ʻāpana o ka genome i hoʻopili māmā ʻia (A compartments, euchromatin) a ʻoi aku ka ikaika o ka transcriptionally, a me nā wahi i paʻa paʻa loa (B compartments, Heterochromatin). Hiki ke hoʻohana ʻia ʻo Hi-C no ke kuhikuhi ʻana i nā Topologically Associated Domains (TADs), nā ʻāpana o ka genome i hoʻopili ʻia nā hale a loaʻa paha nā ʻano hōʻike like, a e ʻike i nā puka lou chromatin, nā ʻāpana DNA i hoʻopaʻa ʻia e nā protein a ʻo ia hoʻi. hoʻonui pinepine ʻia i nā mea hoʻoponopono. Hāʻawi ka lawelawe ʻo Hi-C sequencing ʻo BMKGene i ka poʻe noiʻi e ʻimi i nā ʻāpana spatial o ka genomics, e wehe ana i nā ala hou no ka hoʻomaopopo ʻana i ka hoʻoponopono genome a me kona hopena i ke olakino a me nā maʻi.
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ʻO Chromatin Immunoprecipitation Sequencing (ChIP-seq)
ʻO ka Chromatin Immunoprecipitation (CHIP) kahi hana e hoʻohana ai i nā antibodies e hoʻonui i nā protein paʻa DNA a me kā lākou mau pahuhopu genomics. ʻO kona hoʻohui pū ʻana me NGS e hiki ai i ka genome-wide profiling o nā pahuhopu DNA e pili ana me ka hoʻololi histone, nā kumu transcription, a me nā protein paʻa DNA ʻē aʻe. Hiki i kēia ala hoʻoikaika ke hoʻohālikelike i nā pūnaewele paʻa ma waena o nā ʻano cell like ʻole, nā kiko, a i ʻole nā kūlana. Loaʻa nā noi a ChIP-Seq mai ke aʻo ʻana i ka hoʻoponopono transcriptional a me nā ala hoʻomohala i ka wehewehe ʻana i nā mīkini maʻi, e lilo ia i mea hana pono no ka hoʻomaopopo ʻana i nā ʻāina hoʻoponopono genomic a me ka holomua ʻana i nā ʻike therapeutic.
Papahana: Illumina NovaSeq
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ʻO ke kaʻina hana bisulfite genome holoʻokoʻa (WGBS)
Ke kū nei ʻo Whole Genome Bisulfite Sequencing (WGBS) ma ke ʻano he kumu maʻamau gula no ka ʻimi hohonu ʻana i ka DNA methylation, ʻo ia hoʻi ke kūlana ʻelima i ka cytosine (5-mC), kahi mea hoʻoponopono pivotal o ka hōʻike gene a me ka hana kelepona. ʻO ke kumu o ka WGBS e pili ana i ka lāʻau bisulfite, e hoʻohuli i ka hoʻololi ʻana o nā cytosines unmethylated i uracil (C to U), ʻoiai e waiho ana i nā cytosines methylated i hoʻololi ʻole ʻia. Hāʻawi kēia ʻenehana i ka hoʻonā kumu hoʻokahi, e ʻae i nā mea noiʻi e noiʻi piha i ka methylome a wehe i nā ʻano methylation kūlohelohe e pili ana i nā kūlana like ʻole, ʻo ia hoʻi ka maʻi kanesa. Ma ka hoʻohana ʻana iā WGBS, hiki i nā ʻepekema ke loaʻa nā ʻike like ʻole i nā ʻāina methylation ākea genome, e hāʻawi ana i ka ʻike nuanced o nā mīkini epigenetic e hoʻokumu i nā kaʻina hana olaola like ʻole a me nā maʻi.
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ʻIke no ka Transposase-Accessible Chromatin me ke Kiʻekiʻe Throughput Sequencing (ATAC-seq)
ʻO ATAC-seq kahi hana hoʻokaʻina kiʻekiʻe-throughput i hoʻohana ʻia no ka nānā ʻana i ka genome-wide chromatin accessibility analysis. Hāʻawi ia i ka ʻike hohonu o nā mīkini paʻakikī o ka mana epigenetic honua ma luna o ka hōʻike gen. Ke hoʻohana nei ke ʻano i kahi transposase Tn5 hyperactive e hoʻokaʻawale a hoʻopaʻa i nā ʻāpana chromatin wehe i ka manawa like me ka hoʻokomo ʻana i nā mea hoʻohui. ʻO ka hopena o ka hoʻonui ʻana o PCR i ka hana ʻana i kahi waihona sequencing, e hiki ai ke ʻike piha i nā ʻāpana chromatin hāmama ma lalo o nā kūlana manawa-manawa. Hāʻawi ʻo ATAC-seq i kahi ʻike holoʻokoʻa o nā ʻāina chromatin hiki ke loaʻa, ʻaʻole e like me nā ʻano hana e kālele wale ana i nā kahua paʻa transcription factor a i ʻole nā wahi kikoʻī i hoʻololi ʻia i ka histone. Ma ka hoʻokaʻawale ʻana i kēia mau ʻāpana chromatin wehe, hōʻike ʻo ATAC-seq i nā wahi i ʻoi aku ka nui o nā kaʻina hoʻoponopono hoʻoponopono a me nā pūnaewele hoʻopaʻa transcription factor, e hāʻawi ana i nā ʻike koʻikoʻi i ka hoʻololi ʻana o ka hōʻike gene ma waena o ka genome.
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Hoʻemi ʻia ka hoʻohālikelike Bisulfite Sequencing (RRBS)
Ua puka mai ka Reduced Representation Bisulfite Sequencing (RRBS) ma ke ʻano he kumu kūʻai kūpono a maikaʻi hoʻi i ka Whole Genome Bisulfite Sequencing (WGBS) i ka noiʻi methylation DNA. ʻOiai hāʻawi ʻo WGBS i nā ʻike kikoʻī ma o ka nānā ʻana i ka genome holoʻokoʻa ma ka hoʻonā kumu hoʻokahi, hiki i kāna kumukūʻai kiʻekiʻe ke lilo i mea palena. Hoʻopau maikaʻi ʻo RRBS i kēia pilikia ma ke koho ʻana i kahi ʻāpana o ka genome. Ke hilinaʻi nei kēia ʻano hana i ka hoʻonui ʻia ʻana o nā ʻāina waiwai nui o ka mokupuni ʻo CpG e ka MspI cleavage a ukali ʻia e ka koho nui o nā ʻāpana 200-500/600 bps. No laila, ʻo nā ʻāina kokoke wale i nā mokupuni CpG i hoʻokaʻawale ʻia, ʻoiai ʻo nā mea me nā mokupuni CpG mamao e kāpae ʻia mai ka nānā ʻana. ʻO kēia kaʻina hana, i hui pū ʻia me ka bisulfite sequencing, e ʻae i ka ʻike kiʻekiʻe o ka methylation DNA, a me ke ala hoʻonohonoho, PE150, e nānā pono i nā wēlau o nā mea hoʻokomo ma mua o ka waena, e hoʻonui ana i ka pono o ka methylation profiling. He mea waiwai nui ka RRBS e hiki ai i ka noiʻi DNA methylation ke kumu kūʻai a hoʻonui i ka ʻike o nā mīkini epigenetic.
