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Single-Nuclei RNA Sequencing
Haɓaka kamun cell guda ɗaya da dabarun gina ɗakin karatu na al'ada, haɗe tare da babban tsari, ya kawo sauyi na nazarin maganganun kwayoyin halitta a matakin tantanin halitta. Wannan ci gaban yana ba da damar yin zurfafa da cikakken bincike game da hadaddun yawan adadin tantanin halitta, da shawo kan iyakokin da ke da alaƙa da matsakaicin magana akan dukkan sel da kuma kiyaye ainihin bambance-bambancen tsakanin waɗannan al'ummomi. Yayin da jerin RNA-cell guda ɗaya (scRNA-seq) yana da fa'idodi da ba za a iya musantawa ba, yana fuskantar ƙalubale a wasu kyallen takarda inda ƙirƙirar dakatarwar tantanin halitta guda ɗaya ke da wahala kuma yana buƙatar sabbin samfura. A BMKGene, muna magance wannan ƙalubalen ta hanyar ba da jerin RNA guda ɗaya (snRNA-seq) ta amfani da fasahar zamani ta 10X Genomics Chromium fasaha. Wannan hanya tana faɗaɗa nau'ikan samfuran da za a iya amfani da su don nazarin kwafi a matakin cell-ɗaya.
Keɓewar ƙwayoyin cuta ana samun su ta hanyar sabon guntu na 10X Genomics Chromium, wanda ke nuna tsarin microfluidics na tashoshi takwas tare da ketare biyu. A cikin wannan tsarin, gel beads wanda ya haɗa da barcodes, primers, enzymes, da kuma tsakiya guda ɗaya suna kunshe a cikin nau'in man fetur na nanoliter, suna samar da Gel Bead-in-Emulsion (GEM). Bayan samuwar GEM, sel lysis da sakin lambar sirri suna faruwa a cikin kowane GEM. Daga baya, kwayoyin mRNA suna juyar da rubutu zuwa cDNAs, suna haɗa lambobin barcode 10X da Abubuwan Gano Kwayoyin Halitta na Musamman (UMIs). Waɗannan cDNAs ana sa su zuwa daidaitattun ginin ɗakin karatu, suna sauƙaƙe bincike mai ƙarfi da cikakken bincike na bayanan bayanan kwayoyin halitta a matakin cell-guda.
Platform: 10× Genomics Chromium da Illumina NovaSeq Platform
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10x Genomics Visium Spatial Transcriptome
Fassarar fasikanci fasaha ce mai yanke hukunci wacce ke ba masu bincike damar bincika tsarin maganganun kwayoyin halitta a cikin kyallen takarda yayin da suke kiyaye yanayin sararin samaniya. Ɗayan dandamali mai ƙarfi a cikin wannan yanki shine 10x Genomics Visium haɗe tare da jerin Illumina. Ka'idar 10X Visium ta ta'allaka ne akan guntu na musamman tare da yankin da aka keɓe inda aka sanya sassan nama. Wannan yanki na kama yana ƙunshe da ɓangarorin ɓangarorin, kowanne ya yi daidai da wani wuri na musamman a cikin nama. Kwayoyin RNA da aka kama daga nama ana yi musu lakabi da kebantattun abubuwan gano kwayoyin halitta (UMIs) yayin aiwatar da rubutun baya. Waɗannan tabobin da aka ƙera da UMI suna ba da damar taswirar taswirar sararin samaniya da ƙididdige yawan magana a cikin ƙudurin tantai ɗaya. Haɗuwa da samfuran barcoded na sararin samaniya da UMI suna tabbatar da daidaito da ƙayyadaddun bayanan da aka samar. Ta amfani da wannan fasaha ta Spatial Transcriptomics, masu bincike za su iya samun zurfin fahimta game da tsarin sararin samaniya na sel da hadaddun hulɗar kwayoyin da ke faruwa a cikin kyallen takarda, suna ba da haske mai mahimmanci game da hanyoyin da ke tattare da tsarin ilimin halitta a fannoni da yawa, ciki har da oncology, neuroscience, ilmin halitta na ci gaba, rigakafi. , da kuma nazarin halittu.
Platform: 10X Genomics Visium da Illumina NovaSeq
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Cikakkun Tsawon mRNA Sequencing-Nanopore
Yayin da tsarin mRNA na tushen NGS kayan aiki ne mai dacewa don ƙididdige maganganun kwayoyin halitta, dogaro da gajeriyar karantawa yana taƙaita tasirin sa a cikin hadaddun nazarin kwafin rubutu. A gefe guda, jerin nanopore yana amfani da fasahar dogon karantawa, yana ba da damar jeri cikakken kwafin mRNA. Wannan tsarin yana sauƙaƙe bincike mai zurfi na madadin splicing, gene fusions, poly-adenylation, da ƙididdige abubuwan mRNA.
Tsarin Nanopore, hanyar da ta dogara da siginar lantarki na nanopore guda ɗaya na ainihin lokacin, yana ba da sakamako a cikin ainihin lokaci. Jagoran sunadaran motoci, DNA mai ɗaure biyu yana ɗaure zuwa sunadaran nanopore da aka saka a cikin biofilm, buɗewa yayin da yake wucewa ta tashar nanopore a ƙarƙashin bambancin ƙarfin lantarki. Ana gano siginonin lantarki na musamman waɗanda aka samar ta tushe daban-daban akan madaidaicin DNA kuma ana rarraba su a cikin ainihin lokaci, suna sauƙaƙe daidaitaccen jerin abubuwan nucleotide. Wannan sabuwar dabarar ta shawo kan iyakoki na ɗan gajeren karantawa kuma tana ba da dandamali mai ƙarfi don ƙididdigewar nazarin kwayoyin halitta, gami da hadaddun nazarin fassarar, tare da sakamako nan take.
Dandalin: Nanopore PromethION 48
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Tsarin mRNA mai cikakken tsayi -PacBio
Yayin da tsarin mRNA na tushen NGS kayan aiki ne mai dacewa don ƙididdige maganganun kwayoyin halitta, dogaro da gajeriyar karantawa yana hana amfani da shi a cikin hadaddun nazarin kwafin rubutu. A gefe guda, PacBio sequencing (Iso-Seq) yana amfani da fasahar dogon karantawa, yana ba da damar jeri cikakken kwafin mRNA. Wannan tsarin yana sauƙaƙe bincike mai zurfi na madadin splicing, gene fusions, da poly-adenylation. Koyaya, akwai wasu zaɓuɓɓuka don ƙididdige maganganun kwayoyin halitta saboda yawan adadin bayanan da ake buƙata. Fasahar jeri ta PacBio ta dogara ne akan jerin kwayoyin halitta guda ɗaya, ainihin-lokaci (SMRT), suna ba da fa'ida ta musamman wajen ɗaukar kwafin mRNA mai cikakken tsayi. Wannan sabon tsarin ya ƙunshi amfani da sifili-yanayin waveguides (ZMWs) da microfabricated rijiyoyin da ke ba da damar lura da ainihin lokacin aikin DNA polymerase yayin jerin abubuwa. A cikin waɗannan ZMWs, PacBio's DNA polymerase yana haɗa nau'in DNA na haɗin gwiwa, yana samar da dogon karatu wanda ya kai gabaɗayan kwafin mRNA. Aiki na PacBio a cikin Yanayin Ijma'i na Da'ira (CCS) yana haɓaka daidaito ta hanyar maimaita nau'ikan kwayoyin halitta iri ɗaya. Karatun HiFi da aka ƙirƙira yana da daidaito kwatankwacin NGS, yana ƙara ba da gudummawa ga ingantaccen ingantaccen bincike na fasalulluka masu rikitarwa.
Platform: PacBio Sequel II; PacBio Revio
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Eukaryotic mRNA Sequencing-NGS
Tsarin mRNA, fasaha mai mahimmanci, yana ba da ikon yin cikakken bayanin duk kwafin mRNA a cikin sel a ƙarƙashin takamaiman yanayi. Tare da aikace-aikacen sa mai fa'ida, wannan kayan aiki mai yankewa yana buɗe ƙayyadaddun bayanan bayanan kwayoyin halitta, tsarin halittar, da hanyoyin ƙwayoyin ƙwayoyin cuta masu alaƙa da nau'ikan hanyoyin nazarin halittu. An karɓe shi sosai a cikin bincike na asali, bincike na asibiti, da haɓakar magunguna, jerin mRNA yana ba da haske game da ƙullun tsarin salon salula da tsarin kwayoyin halitta, yana haifar da son sani game da yuwuwar sa a fagage daban-daban.
Platform: Illumina NovaSeq X; DNBSEQ-T7
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Matsakaicin tushen mRNA Sequencing-NGS
Tsarin mRNA yana ba da ikon cikakken bayanin duk bayanan mRNA a cikin sel a ƙarƙashin takamaiman yanayi. Wannan fasaha mai yanke-yanke tana aiki azaman kayan aiki mai ƙarfi, yana buɗe bayanan bayanan ƙwayoyin cuta masu rikitarwa, tsarin halittar halitta, da hanyoyin ƙwayoyin ƙwayoyin cuta masu alaƙa da nau'ikan hanyoyin nazarin halittu. An karɓe shi sosai a cikin bincike na asali, bincike na asibiti, da haɓaka magunguna, jerin mRNA yana ba da haske game da rikitattun kuzarin salon salula da ka'idojin kwayoyin halitta.
Platform: Illumina NovaSeq X; DNBSEQ-T7
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Dogon Ƙididdigar Ƙimar-Illumina
Dogayen RNAs marasa coding (lncRNAs) sun fi nucleotides 200 tsayi waɗanda ke da ƙaramin yuwuwar coding kuma abubuwa ne masu mahimmanci a cikin RNA marasa coding. An samo su a cikin tsakiya da cytoplasm, waɗannan RNAs suna taka muhimmiyar rawa a cikin epigenetic, transcriptional, da ka'idojin rubuce-rubuce, suna nuna mahimmancin su wajen tsara tsarin salula da kwayoyin halitta. Tsarin LncRNA kayan aiki ne mai ƙarfi a cikin bambance-bambancen Cell, Ontogenesis, da cututtukan ɗan adam.
Platform: Illumina NovaSeq
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Ƙananan RNA Sequencing-Illumina
Ƙananan kwayoyin RNA (sRNA), sun haɗa da microRNAs (miRNAs), ƙananan RNAs (siRNAs), da RNAs masu hulɗar piwi (piRNAs). Daga cikin waɗannan, miRNAs, kusan 18-25 nucleotides tsayi, sun fi dacewa musamman don mahimman ayyukansu na tsari a cikin matakai daban-daban na salon salula. Tare da ƙayyadaddun tsarin nama da ƙayyadaddun yanayin magana, miRNAs suna nuna babban kiyayewa a cikin nau'ikan nau'ikan daban-daban.
Platform: Illumina NovaSeq
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Tsarin CircRNA-Illumina
Tsarin RNA madauwari (circRNA-seq) shine yin bayanin martaba da nazarin RNAs madauwari, aji na kwayoyin RNA waɗanda ke samar da rufaffiyar madaukai saboda abubuwan da ba na canonical ba, suna samar da wannan RNA tare da ƙarin kwanciyar hankali. Yayin da aka nuna wasu circRNAs suna aiki azaman soso na microRNA, suna sarrafa microRNAs da hana su daidaita mRNAs ɗin da suke son cimmawa, wasu circRNAs na iya yin hulɗa tare da sunadaran, daidaita maganganun kwayoyin halitta, ko suna da matsayi a cikin tsarin salula. Binciken furuci na circRNA yana ba da haske game da ayyukan gudanarwa na waɗannan ƙwayoyin cuta da mahimmancin su a cikin matakai daban-daban na salon salula, matakan haɓakawa, da yanayin cututtuka, yana ba da gudummawa ga zurfin fahimtar sarƙar ƙa'idar RNA a cikin mahallin maganganun kwayoyin halitta.
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Gabaɗaya Tsarin Rubutu - Illumina
Gabaɗayan jeri na rubutun yana ba da cikakkiyar hanya don bayyana ƙwayoyin RNA daban-daban, wanda ya ƙunshi coding (mRNA) da RNAs mara coding (lncRNA, circRNA, da miRNA). Wannan dabarar tana ɗaukar ɗaukacin rubutun takamaiman sel a wani ɗan lokaci, yana ba da damar cikakkiyar fahimtar hanyoyin salula. Hakanan an san shi da "Jimlar jeri na RNA," yana da nufin buɗe rikitattun hanyoyin sadarwa na tsari a matakin rubutu, yana ba da damar zurfafa bincike kamar gasa RNA endogenous (ceRNA) da kuma nazarin RNA na haɗin gwiwa. Wannan yana nuna matakin farko zuwa ga halayen aiki, musamman a buɗe hanyoyin sadarwar da suka shafi hulɗar ceRNA-miRNA-mRNA tushen ceRNA.
