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Cov khoom

Non-Reference based mRNA Sequencing-NGS

mRNA sequencing txhawb nqa cov kev qhia dav dav ntawm txhua mRNA cov ntawv teev lus hauv cov hlwb hauv cov xwm txheej tshwj xeeb. Qhov kev siv thev naus laus zis no ua haujlwm zoo li cov cuab yeej muaj zog, nthuav tawm cov kab lus sib txawv ntawm cov noob caj noob ces, cov qauv noob, thiab cov txheej txheem molecular cuam tshuam nrog ntau yam txheej txheem lom neeg. Kev siv dav dav hauv kev tshawb fawb, kev kuaj mob, thiab kev tsim tshuaj, mRNA sequencing muab kev nkag siab rau hauv qhov tsis sib xws ntawm cellular dynamics thiab kev tswj caj ces.

Platform: Illumina NovaSeq X; DNBSEQ-T7


Cov ntsiab lus ntawm kev pabcuam

Bioinformatics

Cov txiaj ntsig Demo

Cov ntawv tshaj tawm tshwj xeeb

Nta

● Kev ntes ntawm poly mRNA ua ntej npaj tsev qiv ntawv

● Kev ywj pheej ntawm ib qho kev siv genome: raws li de novo sib sau ua ke ntawm cov ntawv sau tseg, tsim cov npe ntawm cov unigenes uas tau sau tseg nrog ntau lub databases (NR, Swiss-Prot, COG, KOG, eggNOG, Pfam, GO, KEGG)

● Kev tshawb xyuas bioinformatic ntawm cov noob qhia thiab cov qauv sau ntawv

Kev Pabcuam Zoo

Kev txawj ntse: nrog cov ntaub ntawv teev tseg ntawm kev ua tiav ntau dua 600,000 tus qauv ntawm BMKGENE, nthuav dav ntau hom qauv xws li kab lis kev cai ntawm tes, cov ntaub so ntswg, thiab cov kua dej hauv lub cev, peb pab pawg coj ntau yam kev paub rau txhua qhov haujlwm. Peb tau ua tiav kaw 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 lus thiab bioinformatics. Qhov kev soj ntsuam nruj me ntsis ua kom cov khoom xa tuaj raws li cov txiaj ntsig zoo.

● Cov lus piav qhia dav dav: peb siv ntau lub databases los ua haujlwm piav qhia qhov txawv ntawm Cov Genes (DEGs) thiab ua qhov kev txheeb xyuas qhov zoo sib xws, muab kev nkag siab ntawm cov txheej txheem cellular thiab molecular hauv qab cov lus teb.

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 muag kev pabcuam. 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

≥ 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. RNAstable 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

Qauv QC

Kev sim tsim

coj mus kuaj

Qauv xa tuaj

Pilot kev sim

RNA rho tawm

Kev npaj tsev qiv ntawv

Kev tsim kho tsev qiv ntawv

Sequencing

Sequencing

Cov ntaub ntawv tsom xam

Cov ntaub ntawv tsom xam

Kev pab cuam tom qab muag

Kev pab cuam tom qab muag


  • Yav dhau los:
  • Tom ntej:

  • Bioinformatics

    wps_doc_11

    Transscriptome los ua ke thiab xaiv unigene

     

    kev 6

     

     

     Unigene annotation

    xoj 7 

     

    Kev sib raug zoo thiab kev ntsuam xyuas ntawm cov khoom siv roj ntsha

     

     xoj 8

     

     

    Differentially nthuav qhia Genes (DEGs)

     

     kev 9

     

     

    Functional Annotation of DEGs

     

    kev 10

     

    Functional Enrichment of DEGs

     

    kev 11

    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.

     

    Shen, F. et al. (2020) 'De novo transcriptome assembly and sex-biased gene expression in the gonads of Amur catfish (Silurus asotus)', Genomics, 112(3), pp. 2603–2614. doi: 10.1016/J.YGENO.2020.01.026.

    Zhang C., et al. (2016) 'Transcriptome tsom xam ntawm sucrose metabolism thaum lub teeb o thiab kev loj hlob hauv dos (Allium cepa L.),', Frontiers in Plant Science, 7(September), p. 212763. doi: 10.3389/FPLS.2016.01425/BIBTEX.

    Zhou, C. et al. (2017) 'De novo assembly, characterization and annotation for the transcriptome of Sarcoccheilichthys sinensis', PLoS ONE, 12(2). doi: 10.1371/JOURNAL.PONE.0171966.

    Zou, L. et al. (2021) 'De novo transcriptome tsom xam muab kev nkag siab rau cov ntsev kam rau ntawm Podocarpus macrophyllus nyob rau hauv salinity stress', BMC Plant Biology, 21(1), pp. 1–17. doi: 10.1186/S12870-021-03274-1/FIGURES/9.

    tau ib nqe lus

    Sau koj cov lus ntawm no thiab xa tuaj rau peb

    Xa koj cov lus rau peb: