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Ke Kaʻina Metagenomic -NGS
ʻO ka metagenome kahi hōʻiliʻili o ka nui o nā mea genetic o kahi kaiāulu hui o nā meaola, e like me nā metagenomes kaiapuni a me ke kanaka. Loaʻa iā ia nā genomes o nā microorganisms hiki ke kanu a kanu ʻole ʻia. Hiki i ka Shotgun metagenomic sequencing me NGS ke aʻo ʻana i kēia mau ʻāina genomic paʻakikī i hoʻokomo ʻia i loko o nā laʻana kaiapuni ma o ka hāʻawi ʻana i nā mea ʻoi aku ma mua o ka taxonomic profiling, me ka hāʻawi ʻana i nā ʻike kikoʻī i nā ʻano ʻano like ʻole, ka nui o ka dynamics, a me nā hale paʻakikī. Ma waho aʻe o nā haʻawina ʻauhau, hāʻawi pū ka metagenomics shotgun i kahi hiʻohiʻona genomics hana, hiki ke ʻimi i nā ʻano genes i hoʻopaʻa ʻia a me kā lākou hana putative i nā kaʻina kaiaola. ʻO ka mea hope loa, ʻo ka hoʻokumu ʻana i nā ʻupena hoʻopili ma waena o nā mea genetic a me nā mea pili i ke kaiapuni e hāʻawi i ka ʻike holoʻokoʻa o ka pilina paʻakikī ma waena o nā kaiāulu microbial a me ko lākou ʻano kaiaola. I ka hopena, kū ka metagenomic sequencing ma ke ʻano he mea nui no ka wehe ʻana i nā paʻakikī genomic o nā kaiāulu microbial like ʻole, e hoʻomālamalama ana i nā pilina multifaceted ma waena o ka genetics a me ka ecology i loko o kēia mau kaiaola paʻakikī.
Nā papahana: Illumina NovaSeq a me DNBSEQ-T7
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Hoʻokaʻina Metagenomic-TGS
ʻO ka metagenome kahi hōʻiliʻili o nā mea genetic o kahi hui hui o nā mea ola, e like me nā metagenomes kaiapuni a me ke kanaka. Loaʻa iā ia nā genomes o nā microorganisms hiki ke kanu a kanu ʻole ʻia. Hiki i ka metagenomic sequencing ke aʻo ʻana i kēia mau ʻāina genomic paʻakikī i hoʻokomo ʻia i loko o nā laʻana kaiaola ma o ka hāʻawi ʻana i nā mea ʻoi aku ma mua o ka taxonomic profiling. Hāʻawi pū ia i kahi hiʻohiʻona genomics hana ma ka ʻimi ʻana i nā genes i hoʻopili ʻia a me kā lākou hana putative i nā kaʻina kaiapuni. ʻOiai ua hoʻohana nui ʻia ka hoʻokokoke ʻana o ka pana pana kuʻuna me ka hoʻopili ʻana o Illumina i nā haʻawina metagenomic, ʻo ka hiki ʻana mai o Nanopore a me PacBio ka lōʻihi heluhelu ʻana ua hoʻololi i ke kahua. Hoʻonui ka ʻenehana Nanopore a me PacBio i nā loiloi bioinformatic i lalo, ʻo ia hoʻi ka hui metagenome, e hōʻoiaʻiʻo ana i nā hui hoʻomau. Hōʻike nā hōʻike ua hoʻokumu maikaʻi nā metagenomics i hoʻokumu ʻia ʻo Nanopore a me PacBio i nā genomes bacterial piha a pani ʻia mai nā microbiomes paʻakikī (Moss, EL, et al., Nature Biotech, 2020). ʻO ka hoʻohui ʻana i ka heluhelu ʻana o Nanopore me ka heluhelu ʻana o Illumina e hāʻawi i kahi ala hoʻolālā no ka hoʻoponopono hewa, e hoʻemi ana i ka pololei haʻahaʻa haʻahaʻa o Nanopore. Hoʻohana kēia hui synergistic i nā ikaika o kēlā me kēia kahua hoʻonohonoho, e hāʻawi ana i kahi hopena paʻa e lanakila ai i nā palena kūpono a me ka holomua ʻana i ka pololei a me ka hilinaʻi o nā loiloi metagenomic.
Papahana: Nanopore PromethION 48, Illumia a me PacBio Revio
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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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16S/18S/ITS Amplicon Sequencing-PacBio
ʻO nā genes 16S a me 18S rRNA, me ka ʻāpana Internal Transcribed Spacer (ITS), e lilo i mau māka manamana lima molecular pivotal ma muli o kā lākou hui pū ʻana o nā ʻāpana i mālama nui ʻia a hyper-variable, e hoʻolilo iā lākou i mea waiwai nui no ka hoʻohālikelike ʻana i nā meaola prokaryotic a me eukaryotic. Hāʻawi ka hoʻonui a me ka hoʻonohonoho ʻana o kēia mau wahi i kahi ala kaʻawale ʻole no ka noiʻi ʻana i ka haku mele microbial a me nā ʻokoʻa ma waena o nā ʻano kaiaola. ʻOiai ʻo Illumina sequencing maʻamau e hoʻopaʻa i nā wahi hypervariable pōkole e like me V3-V4 o 16S a me ITS1, ua hōʻike ʻia e hiki ke hoʻokō ʻia ka annotation taxonomic kiʻekiʻe ma ke kaʻina ʻana i ka lōʻihi piha o 16S, 18S, a me ITS. Loaʻa i kēia ʻano hoʻokō piha i nā pākēneka kiʻekiʻe o nā kaʻina i hoʻokaʻawale pololei ʻia, e loaʻa ana i kahi pae o ka hoʻonā e hoʻonui i ka ʻike ʻano. Kūʻē ʻo PacBio's Single-Molecule Real-Time (SMRT) sequencing platform ma ka hāʻawi ʻana i nā heluhelu lōʻihi loa (HiFi) e uhi ana i nā amplicons lōʻihi piha, e hoʻokūkū ana i ka pololei o ke kaʻina Illumina. ʻO kēia hiki ke hiki i nā mea noiʻi ke loaʻa kahi pōmaikaʻi like ʻole - kahi hiʻohiʻona panoramic o ka ʻāina genetic. Hoʻokiʻekiʻe nui ʻia ka uhi ʻana i ka hoʻonā ʻana i ka moʻolelo anoano, i loko o nā kaiāulu bacteria a fungal paha, e hiki ai ke ʻike hohonu i ka paʻakikī o nā heluna microbial.
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16S/18S/ITS Amplicon Sequencing-NGS
ʻO ka hoʻopili ʻana o Amplicon me ka ʻenehana Illumina, e kuhikuhi pono ana i ka 16S, 18S, a me ITS genetic markers, he ala ikaika no ka wehe ʻana i ka phylogeny, taxonomy, a me ka nui o nā ʻano i loko o nā kaiāulu microbial. ʻO kēia ala e pili ana i ka hoʻonohonoho ʻana i nā ʻāpana hypervariable o nā hōʻailona genetic housekeeping. Hoʻokomo mua ʻia ma ke ʻano he molecular fingerprint eWoeses et ali ka makahiki 1977, ua hoʻololi kēia ʻenehana i ka microbiome profiling ma o ka ʻae ʻana i nā loiloi kaʻawale ʻole. Ma o ke kaʻina ʻana o 16S (bacteria), 18S (fungi), a me Internal Transcribed Spacer (ITS, fungi), hiki i nā mea noiʻi ke ʻike ʻaʻole wale i nā ʻano nui wale nō akā i nā mea ʻike ʻole a ʻike ʻole ʻia. Hoʻohana nui ʻia ma ke ʻano he mea hana pivotal, ua lilo ka amplicon sequencing i mea kōkua i ka ʻike ʻana i nā haku mele microbial ʻokoʻa ma waena o nā kaiapuni like ʻole, me ka waha o ke kanaka, ka ʻōpū, ka stool, a ma waho.
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ʻO ka hoʻopaʻa ʻana i ka genome holoʻokoʻa i ka maʻi bacteria a me ka fungal
He mea koʻikoʻi nā papahana hoʻonohonoho hou-genome holoʻokoʻa a me ka fungal no ka holomua ʻana i nā genomics microbial ma o ka hiki ke hoʻopau a me ka hoʻohālikelike ʻana i nā genome microbial. Hoʻomaʻamaʻa kēia i ka ʻenekinia fermentation, ka hoʻonui ʻana i nā kaʻina hana ʻoihana, a me ka ʻimi ʻana i nā ala metabolism lua. Eia kekahi, he mea koʻikoʻi ka hoʻopili hou ʻana o ka fungal a me ka bacteria no ka hoʻomaopopo ʻana i ka hoʻololi ʻana i ke kaiapuni, ka hoʻopaʻa ʻana i nā ʻano, a me ka hōʻike ʻana i ka hoʻomohala genetic evolution, me nā hopena ākea i ka lāʻau lapaʻau, ka mahiʻai, a me ka ʻepekema kaiapuni.
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PacBio-Ka lōʻihi piha 16S/18S/ITS Amplicon Sequencing
Hoʻokumu ʻia ka paepae Amplicon (16S/18S/ITS) me nā makahiki o ka ʻike ʻana i ka noiʻi papahana microbial diversity, kahi i hoʻopaʻa ʻia i ka nānā ʻana maʻamau a me ka loiloi pilikino: uhi ʻia ka loiloi kumu i ka ʻike loiloi kumu o ka noiʻi microbial o kēia manawa, waiwai a piha ka ʻikepili. a hōʻike ʻia nā hopena o ka nānā ʻana ma ke ʻano o nā hōʻike papahana; He ʻokoʻa ka ʻike o ka loiloi pilikino. Hiki ke koho ʻia nā laʻana a hiki ke hoʻonohonoho pono ʻia nā ʻāpana e like me ka hōʻike loiloi kumu a me ke kumu noiʻi, e ʻike ai i nā koi pilikino. ʻO ka ʻōnaehana hana Windows, maʻalahi a wikiwiki.
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PacBio-Full-lōʻihi Transcriptome (Non-Reference)
Ke lawe nei i ka ʻikepili hoʻokaʻina Isoform i ka Pacific Biosciences (PacBio) ma ke ʻano he hoʻokomo, hiki i kēia App ke ʻike i nā kaʻina transcript lōʻihi (me ka ʻole o ka hui ʻana). Ma ka palapala ʻana i nā kaʻina holoʻokoʻa kūʻē i ka genome kuhikuhi, hiki ke hoʻopaʻa ʻia nā transcripts e nā genes i ʻike ʻia, transcripts, coding regions, etc. ʻO ka loiloi hui pū me ka NGS transcriptome sequencing data e hiki ai i ka hōʻike kikoʻī a me ka helu pololei ʻana i ka hōʻike ma ka pae transcript, ka mea e pōmaikaʻi nui ai i ka hōʻike ʻokoʻa a me ka nānā ʻana i ka hana.
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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.
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Ka hoʻokaʻina RNA Prokaryotic
Hāʻawi ka RNA sequencing i ka hōʻike piha ʻana o nā transcript RNA āpau i loko o nā cell ma lalo o nā kūlana kikoʻī. Ke hana nei kēia ʻenehana ʻokiʻoki ma ke ʻano he mea hana ikaika, e wehe ana i nā moʻolelo hōʻikeʻike gene paʻakikī, nā ʻano hana gene, a me nā mīkini molekala e pili ana i nā kaʻina hana olaola like ʻole. Hoʻohana nui ʻia i ka noiʻi koʻikoʻi, nā diagnostics clinical, a me ka hoʻomohala ʻana i ka lāʻau lapaʻau, hāʻawi ʻo RNA sequencing i nā ʻike i loko o ka intricacies o ka dynamics cellular a me ka hoʻoponopono genetic. Hoʻopili ʻia kā mākou prokaryotic RNA sample process no prokaryotic transcriptomes, e pili ana i ka rRNA depletion a me ka hoʻomākaukau ʻana i ka waihona kuhikuhi.
Papahana: Illumina NovaSeq
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Hoʻokaʻina Metatranscriptome
Me ka hoʻohana ʻana i ka ʻenehana sequencing Illumina, ʻo ka BMKGENE's metatranscriptome sequencing service e hōʻike ana i ka hōʻike gene dynamic o nā ʻano microbes like ʻole, e holo ana i nā eukaryotes i nā prokaryotes a me nā maʻi maʻi, i loko o nā kaiapuni kūlohelohe e like me ka lepo, ka wai, ke kai, ka ʻōpū, a me ka ʻōpū. Hāʻawi kā mākou lawelawe holoʻokoʻa i ka poʻe noiʻi e ʻimi i nā moʻolelo hōʻike gene piha o nā kaiāulu microbial. Ma waho aʻe o ka loiloi taxonomic, hoʻomaʻamaʻa kā mākou lawelawe ʻoihana metatranscriptome i ka ʻimi ʻana i ka hoʻonui ʻana i ka hana, e hoʻomālamalama ana i nā genes i hōʻike ʻia a me kā lākou mau kuleana. Wehe i ka waiwai o nā ʻike olaola i kou hoʻokele ʻana i nā ʻāina paʻakikī o ka hōʻike gene, ka ʻokoʻa ʻauhau, a me nā hana hoʻoikaika i loko o kēia mau niches kaiapuni.
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De novo Fungal Genome Assembly
Hāʻawi ʻo BMKGENE i nā hoʻonā maʻalahi no nā genomes fungal, e mālama ana i nā pono noiʻi like ʻole a me ka piha genome makemake. ʻO ka hoʻohana ʻana i ke kaʻina Illumina heluhelu pōkole wale nō e hiki ai ke hana i kahi kikoʻī genome. Hoʻohui ʻia nā heluhelu pōkole a me ka heluhelu lōʻihi me Nanopore a i ʻole Pacbio no kahi genome fungal i hoʻomaʻemaʻe ʻia me nā contigs lōʻihi. Eia kekahi, ʻo ka hoʻohui ʻana i ka hi-C sequencing e hoʻonui hou i nā hiki, e hiki ai ke loaʻa i kahi genome pae chromosome piha.
