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Ukulandelana kwe-Metagenomic -NGS
I-metagenome iqoqo lezinto eziphathelene nezakhi zofuzo zomphakathi oxubile wezinto eziphilayo, njenge-metagenome yezemvelo neyabantu. Iqukethe ama-genome ezinto ezincane ezingatshalwa kanye nezingalinywa. Ukulandelana kwe-metagenomic nge-Shotgun nge-NGS kwenza kube lula ukutadisha lezi zindawo eziyinkimbinkimbi ze-genomic ezifakwe kumasampula ezemvelo ngokunikeza okungaphezu kokulandelelana kwe-taxonomic, okunikeza nokuqonda okuningiliziwe ngokuhlukahluka kwezinhlobo, ukuguquguquka kobuningi, kanye nezakhiwo eziyinkimbinkimbi zabantu. Ngaphandle kwezifundo ze-taxonomic, i-shotgun metagenomics iphinde inikeze umbono osebenzayo we-genomics, okuvumela ukuhlola izakhi zofuzo ezifakiwe kanye nezindima zazo ezibeka engcupheni yezemvelo. Okokugcina, ukusungulwa kwamanethiwekhi okuxhumana phakathi kwezakhi zofuzo kanye nezici zemvelo kunikela ekuqondeni okuphelele kokuxhumana okuyinkimbinkimbi phakathi kwemiphakathi yamagciwane kanye nesizinda sayo semvelo. Ekuphetheni, ukulandelana kwe-metagenomic kumi njengethuluzi elibalulekile lokwembula ubunzima be-genomic bemiphakathi yamagciwane ahlukahlukene, kukhanyise ubudlelwano obunezinhlangothi eziningi phakathi kwezakhi zofuzo kanye nemvelo ngaphakathi kwalezi zindawo zemvelo eziyinkimbinkimbi.
Amapulatifomu: Illumina NovaSeq kanye ne-DNBSEQ-T7
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Ukulandelana kwe-Metagenomic-TGS
I-metagenome iqoqo lezinto zofuzo zomphakathi oxubile wezinto eziphilayo, njenge-metagenome yezemvelo neyabantu. Iqukethe ama-genome ezinto ezincane ezitshalwa kanye nezingalinywa. Ukulandelana kwe-metagenomic kwenza ucwaningo lwalezi zindawo eziyinkimbinkimbi ze-genomic ezifakwe kumasampula emvelo ngokunikeza okungaphezu kokulandelela i-taxonomic. Iphinde inikeze umbono osebenzayo we-genomics ngokuhlola izakhi zofuzo ezifakiwe kanye nezindima zazo ezibeka engcupheni yezemvelo. Ngenkathi izindlela zendabuko ze-shotgun nge-Illumina sequencing zisetshenziswe kabanzi ezifundweni ze-metagenomic, ukufika kokulandelana kwe-Nanopore ne-PacBio okufundwa isikhathi eside kushintshe insimu. Ubuchwepheshe be-Nanopore ne-PacBio buthuthukisa ukuhlaziywa kwe-bioinformatic okuphansi, ikakhulukazi ukuhlanganiswa kwe-metagenome, ukuqinisekisa ukuhlanganiswa okuqhubekayo. Imibiko ikhombisa ukuthi ama-metagenomics asekelwe ku-Nanopore kanye ne-PacBio akhiqize ngempumelelo ama-genome aphelele futhi avaliwe avela kuma-microbiomes ayinkimbinkimbi (Moss, EL, et al., Nature Biotech, 2020). Ukuhlanganisa ukufundwa kwe-Nanopore nokufundwa kwe-Illumina kunikeza indlela ehlakaniphile yokulungisa amaphutha, kunciphisa ukunemba okuphansi kwe-Nanopore. Le nhlanganisela yokusebenzisana isebenzisa amandla epulatifomu ngayinye yokulandelana, inikeza ikhambi eliqinile lokunqoba imikhawulo engaba khona kanye nokuthuthukisa ukunemba nokuthembeka kokuhlaziywa kwe-metagenomic.
Ipulatifomu: I-Nanopore PromethION 48, i-Illumia kanye ne-PacBio Revio
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16S/18S/ITS Amplicon Sequencing-PacBio
Izakhi zofuzo ze-rRNA ze-16S kanye ne-18S, kanye nesifunda se-Internal Transcribed Spacer (ITS), zisebenza njengezimpawu ezibalulekile zokubhala iminwe yama-molecule ngenxa yokuhlanganiswa kwazo kwezifunda ezigcinwe kakhulu neziguquguqukayo kakhulu, okwenza zibe amathuluzi abalulekile okuhlukanisa izinto eziphilayo ze-prokaryotic kanye ne-eukaryotic. Ukukhulisa nokulandelana kwalezi zifunda kunikeza indlela engenakho ukuhlukaniswa yokuphenya ukwakheka kwamagciwane kanye nokwehlukahluka kuzo zonke izindawo zemvelo ezahlukahlukene. Ngenkathi ukulandelana kwe-Illumina kuvame ukuqondisa izifunda ezimfushane eziguquguqukayo njenge-V3-V4 ye-16S kanye ne-ITS1, kuye kwabonakala ukuthi isichasiselo se-taxonomic esiphezulu singatholakala ngokulandelana ubude obugcwele be-16S, 18S, kanye ne-ITS. Le ndlela ephelele iholela kumaphesenti aphezulu okulandelana okuhlukaniswe ngokunembile, kufinyelelwa izinga lokuxazululwa elifinyelela ekuhlonzweni kwezinhlobo. Ipulatifomu yokulandelelana kwe-Single-Molecule Real-Time (SMRT) ye-PacBio ivelele ngokunikeza ukufundwa okude okunembe kakhulu (i-HiFi) okuhlanganisa ama-amplicons agcwele ubude, okuncintisana nokunemba kokulandelana kwe-Illumina. Leli khono livumela abacwaningi ukuthi bathole inzuzo engenakuqhathaniswa - umbono obanzi wendawo yezakhi zofuzo. Ukumbozwa okunwetshiwe kuphakamisa kakhulu isinqumo ekuchazweni kwezinhlobo, ikakhulukazi emiphakathini yamagciwane noma yesikhunta, okwenza kube nokuqonda okujulile kobunzima bezinhlobo zamagciwane.
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16S/18S/ITS Ukulandelelana kwe-Amplicon-NGS
Ukulandelana kwe-Amplicon ngobuchwepheshe be-Illumina, okuqondiswe ngqo kuma-genetic markers e-16S, 18S, kanye ne-ITS, kuyindlela enamandla yokwembula i-phylogeny, i-taxonomy, kanye nobuningi bezinhlobo emiphakathini yamagciwane. Le ndlela ihilela ukulandelana kwezindawo eziguquguqukayo kakhulu zama-genetic markers. Ekuqaleni yethulwa njengophawu lweminwe lwama-molecule yiUWoeses nabanyeNgo-1977, le ndlela ishintshe kakhulu ukuqoqwa kwe-microbiome ngokuvumela ukuhlaziywa okungenazo izitho zomuntu. Ngokulandelana kwe-16S (amabhaktheriya), i-18S (isikhunta), kanye ne-Internal Transcribed Spacer (ITS, isikhunta), abacwaningi abakwazi ukubona izinhlobo eziningi kuphela kodwa futhi nezinhlobo ezingavamile nezingaziwa. Njengoba yamukelwa kabanzi njengethuluzi elibalulekile, ukulandelana kwe-amplicon kuye kwaba usizo ekuqondeni ukwakheka okuhlukile kwamagciwane ezindaweni ezahlukahlukene, kufaka phakathi umlomo womuntu, amathumbu, indle, nangale kwalokho.
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Ukuhlelwa Kabusha Kwe-Genome Ephelele Yama-bacteria Ne-Fungal
Amaphrojekthi okulandelana kabusha kwezakhi zofuzo eziphelele zamagciwane kanye nesikhunta abalulekile ekuthuthukisweni kwezakhi zofuzo ezibangelwa amagciwane ngokwenza kube lula ukuqedwa nokuqhathaniswa kwezakhi zofuzo ezibangelwa amagciwane. Lokhu kwenza kube lula ubunjiniyela bokuvubela, ukwenza ngcono izinqubo zezimboni, kanye nokuhlola izindlela zesibili zemetabolism. Ngaphezu kwalokho, ukulandelana kabusha kwezakhi zofuzo kanye nebhaktheriya kubalulekile ekuqondeni ukuzivumelanisa nemvelo, ukuthuthukisa izinhlobo, kanye nokwembula ukuguquguquka kwezakhi zofuzo, okunemiphumela ebanzi kwezokwelapha, ezolimo, kanye nesayensi yezemvelo.
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Ukulandelana kwe-Prokaryotic RNA
Ukulandelana kwe-RNA kunika amandla ukuqoshwa okuphelele kwazo zonke izinguqulo ze-RNA ngaphakathi kwamaseli ngaphansi kwezimo ezithile. Lobu buchwepheshe obusezingeni eliphezulu busebenza njengethuluzi elinamandla, lembula amaphrofayili okubonakaliswa kwezakhi zofuzo ayinkimbinkimbi, izakhiwo zezakhi zofuzo, kanye nezinqubo zama-molecule ezihlotshaniswa nezinqubo ezahlukahlukene zebhayoloji. Njengoba kusetshenziswa kabanzi ocwaningweni oluyisisekelo, ekuxilongweni kwezokwelapha, kanye nasekuthuthukisweni kwemithi, ukulandelana kwe-RNA kunikeza ukuqonda ngobunzima be-dynamics yamaseli kanye nokulawulwa kwezakhi zofuzo. Ukucubungula kwethu isampula ye-prokaryotic RNA kwenzelwe ama-transcriptome e-prokaryotic, okubandakanya ukuncipha kwe-rRNA kanye nokulungiselela umtapo wolwazi oqondisayo.
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Ukulandelana kwe-Metatranscriptome
Isebenzisa ubuchwepheshe bokulandelana kwe-Illumina, insizakalo yokulandelana kwe-metatranscriptome ye-BMKGENE yembula ukubonakaliswa kwezakhi zofuzo okuguquguqukayo kwezinhlobo ezahlukene zamagciwane, okusabalala kuma-eukaryotes kuya kuma-prokaryotes namagciwane, ngaphakathi kwezindawo zemvelo ezifana nomhlabathi, amanzi, ulwandle, indle, kanye namathumbu. Insizakalo yethu ephelele inika amandla abacwaningi ukuthi bahlole amaphrofayili aphelele okubonakaliswa kwezakhi zofuzo emiphakathini yamagciwane. Ngaphandle kokuhlaziywa kwe-taxonomic, insizakalo yethu yokulandelana kwe-metatranscriptome yenza kube lula ukuhlola ukucebisa okusebenzayo, okukhanyisa izakhi zofuzo ezivezwe ngokuhlukile kanye nezindima zazo. Thola ulwazi oluningi lwezinto eziphilayo njengoba uhamba ezindaweni eziyinkimbinkimbi zokubonakaliswa kwezakhi zofuzo, ukuhlukahluka kwe-taxonomic, kanye nokuguquguquka kokusebenza ngaphakathi kwalezi zindawo zemvelo ezahlukahlukene.
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I-Novo Fungal Genome Assembly
I-BMKGENE inikeza izixazululo ezihlukahlukene zama-genome esikhunta, ihlangabezana nezidingo zocwaningo ezahlukahlukene kanye nokuphelela kwe-genome okufiswayo. Ukusebenzisa i-short-read Illumina sequencing kuphela kuvumela ukukhiqizwa kwe-draft genome. I-short-reading kanye ne-long-read sequencing kusetshenziswa i-Nanopore noma i-Pacbio kuhlanganiswe ukuze kube ne-fungus genome ecwengekile kakhulu enezinhlayiya ezinde.
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I-Novo Bacterial Genome Assembly
Sinikeza isevisi ephelele yokuhlanganisa i-genome yamagciwane, okuqinisekisa izikhala ezingu-0. Lokhu kungenzeka ngokuhlanganisa ubuchwepheshe bokulandelana kokufunda isikhathi eside, njenge-Nanopore ne-PacBio yokuhlanganisa nokulandelana kokufunda okufushane ne-Illumina yokuqinisekisa ukuhlanganiswa nokulungiswa kwamaphutha okufundwa kwe-ONT. Insizakalo yethu inikeza ukuhamba komsebenzi okuphelele kwe-bioinformatic kusukela ekuhlanganisweni, isichasiselo esisebenzayo, kanye nokuhlaziywa kwe-bioinformatic okuthuthukisiwe, kugcwalisa imigomo ethile yocwaningo. Le nsizakalo ivumela ukuthuthukiswa kwe-genome eqondile yezifundo ezahlukene ze-genetic kanye ne-genomic. Ngaphezu kwalokho, yakha isisekelo sezinhlelo zokusebenza ezifana nokwenza ngcono ukucindezeleka, ubunjiniyela be-genetic, kanye nokuthuthukiswa kobuchwepheshe be-microbial, ukuqinisekisa idatha ye-genomic ethembekile nengenazikhala ebalulekile ekuthuthukiseni ukuqonda kwesayensi kanye nokusungula izinto ezintsha ze-biotechnology.
