Eukaryotic mRNA Sequencing-NGS
Nta
● Kev ntes ntawm poly mRNA ua ntej kev npaj tsev qiv ntawv
● Kev xaiv cov kev taw qhia mRNA cov tsev qiv ntawv npaj kom tau txais cov ntaub ntawv sib txuas tshwj xeeb
● Bioinformatic tsom xam ntawm cov noob qhia thiab cov qauv sau ntawv
Qhov zoo
●Kev txawj ntse: Peb pab pawg tau ua tiav ntau dua 600,000 tus qauv ntawm BMKgene, nthuav tawm ntau hom qauv xws li kab lis kev cai ntawm tes, cov ntaub so ntswg, thiab cov kua dej hauv lub cev. Peb nqa cov kev paub dhau los rau txhua qhov haujlwm thiab tau ua tiav ntau dua 100,000 mRNA-Seq cov haujlwm hauv ntau qhov kev tshawb fawb.
●Rigorous Quality Control: Peb siv cov ntsiab lus tseem ceeb hauv txhua theem, los ntawm cov qauv thiab kev npaj tsev qiv ntawv mus rau kev sib txuas thiab bioinformatics. Qhov kev soj ntsuam xyuas zoo no ua kom cov khoom xa tuaj raws li cov txiaj ntsig zoo, ua rau koj paub tseeb tias koj qhov project yog nyob rau hauv kev ntseeg siab.
●Kev piav qhia dav: Peb siv ntau lub chaw khaws ntaub ntawv los ua haujlwm piav qhia qhov txawv ntawm Cov Genes (DEGs) thiab ua cov kev txheeb xyuas cov txiaj ntsig zoo sib xws. Qhov kev qhia dav dav no muab kev nkag siab txog cov txheej txheem ntawm tes thiab cov txheej txheem molecular hauv qab cov lus teb, kom ntseeg tau tias koj tau txais kev paub txog koj qhov project cov txiaj ntsig.
●Kev them nyiaj yug tom qab muag: Peb qhov kev cog lus txuas ntxiv dhau qhov kev ua tiav nrog rau 3 lub hlis tom qab kev muag khoom. Nyob rau lub sijhawm no, peb muab cov phiaj xwm taug qab, kev pab daws teeb meem, thiab Q&A ntu los daws cov lus nug ntsig txog cov txiaj ntsig.
Sample Requirements thiab Delivery
| Lub tsev qiv ntawv | Sequencing tswv yim | Cov ntaub ntawv pom zoo | Kev Tswj Xyuas Zoo |
| Poly A enriched | Illumina PE 150 DNBSEQ-T7 | 6-10 Gb | Q30≥85% |
Cov Qauv Xav Tau:
Nucleotides:
| Conc.(ng/μl) | Tus nqi (μg) | Purity | Kev ncaj ncees | |
| Standard Library | ≥ 10 | ≥ 0.2 | OD 260/280 = 1.7-2.5 OD 260/230 = 0.5-2.5 Txwv los yog tsis muaj protein ntau lossis DNA kis tau pom ntawm gel. | Rau cov nroj tsuag: RIN≥4.0; Rau tsiaj: RIN≥4.5; 5.0 ≥28S / 18S≥1.0; txwv los yog tsis muaj qhov nce siab |
| Directional Library | ≥ 10 | ≥ 0.2 | OD 260/280 = 1.7-2.5 OD 260/230 = 0.5-2.5 Txwv los yog tsis muaj protein ntau lossis DNA kis tau pom ntawm gel. | Rau cov nroj tsuag: RIN≥4.0; Rau tsiaj: RIN≥4.5; 5.0 ≥28S / 18S≥1.0; txwv los yog tsis muaj qhov nce siab |
● Nroj tsuag:
Hauv paus, qia los yog Petal: 450 mg
Nplooj los yog noob: 300 mg
Txiv Hmab Txiv Ntoo: 1.2 g
●Tsiaj:
Lub plawv los yog txoj hnyuv: 300 mg
Viscera lossis hlwb: 240 mg
Cov leeg: 450 mg
Pob txha, plaub hau lossis tawv nqaij: 1g
● Arthropods:
qog: 6g
Crustacea: 300 mg
● Tag nrho cov ntshav:1 cev
● Cells: 106 hlwb
Pom zoo coj mus kuaj
Ntim: 2 ml centrifuge tube (Tin foil tsis pom zoo)
Sample labeling: Pab pawg + replicate xws li A1, A2, A3; B1, B2, B3....
Kev xa khoom:
1. Dry-ice: Cov qauv yuav tsum tau ntim rau hauv hnab thiab faus rau hauv cov dej khov qhuav.
2. RNAs lub rooj raj: RNA cov qauv tuaj yeem qhuav hauv RNA stabilization raj (xws li RNAstable®) thiab xa mus rau hauv chav tsev kub.
Kev Pabcuam Ua Haujlwm Flow
Kev sim tsim
Qauv xa tuaj
RNA rho tawm
Kev tsim kho tsev qiv ntawv
Sequencing
Cov ntaub ntawv tsom xam
Kev pab cuam tom qab muag
Bioinformatics
Eukaryotic mRNA sequencing tsom xam workflow
Bioinformatics
ØRaw cov ntaub ntawv zoo tswj
ØReference genome alignment
ØTranscript structure analysis
ØKev nthuav qhia kom muaj nuj nqis
ØKev txheeb xyuas qhov sib txawv
ØFunction annotation thiab enrichment
Siv Genome Alignment
Cov ntaub ntawv Saturation
Kev sib raug zoo thiab kev ntsuam xyuas ntawm cov khoom siv roj ntsha
Differentially nthuav qhia Genes (DEGs)
Functional Annotation of DEGs
Functional Enrichment of DEGs
Tshawb nrhiav cov kev nce qib uas tau pab los ntawm BMKGene's eukaryotic NGS mRNA sequencing cov kev pabcuam los ntawm kev sau cov ntawv tshaj tawm.
Huang, L. et al. (2023) 'Triclosan thiab triclocarban tsis muaj zog lub peev xwm ntawm cov ntses kub los ntawm kev txwv cov ntxhiab tsw, cuam tshuam olfactory teeb liab, thiab cuam tshuam olfactory cov ntaub ntawv ua', Dej Tshawb Fawb, 233, p. 119736. doi: 10.1016/J.WATRES.2023.119736.
Jia, LJ et al. (2023) 'Aspergillus fumigatus hijacks human p11 to redirect fungal-muaj phagosomes to non-degradative pathway', Cell Host & Microbe, 31(3), pp. 373-388.e10. doi: 10.1016/J.CHOM.2023.02.002.
Jin, K. et al. (2022) 'TCP Transcription Factors Involved in Shoot Development of Ma Bamboo (Dendrocalamus latiflorus Munro),', Frontiers in Plant Science, 13, p. 884443. doi: 10.3389/FPLS.2022.884443/BIBTEX.
Wen, X. et al. (2022) 'Chrysanthemum lavandulifolium genome thiab molecular mechanism nyob rau hauv ntau hom capitulum', Horticulture Research, 9. doi: 10.1093/HR/UHAB022.
Zhang, Yujie et al. (2023) 'A cascade nanoreactor rau kev txhim kho kev kho sonodynamic ntawm mob qog nqaij hlav hauv plab los ntawm kev sib koom ua ke ROS augment thiab autophagy blockage', Nano Hnub no, 49, p. 101798. doi: 10.1016/J.NANTOD.2023.101798.
