Ntev Tsis-coding Sequencing-Illumina
Kev Pabcuam Zoo
●Kev sib koom ua ke ntawm mRNA thiab lncRNA: los ntawm kev sib txuas cov lej ntawm mRNA cov ntawv sau tseg nrog kev kawm ntawm lncRNA thiab lawv lub hom phiaj, nws muaj peev xwm kom tau txais qhov tob tob ntawm cov txheej txheem tswj hwm hauv qab ntawm cov lus teb ntawm tes.
●Kev txawj ntse: Peb tau ua tiav ntau dua 230,000 tus qauv, nthuav dav cov qauv sib txawv thiab cov phiaj xwm 'lub hom phiaj. Peb coj kev nplua nuj ntawm kev txawj ntse rau txhua qhov project.
●Kev sib koom ua ke ntawm mRNA thiab lncRNA: Peb muab qhov ntau ntawm mRNA cov ntaub ntawv sau tseg nrog kev kawm ntawm lncRNA thiab lawv cov hom phiaj, ua kom nws muaj peev xwm kom tau txais cov ntsiab lus tob ntawm cov txheej txheem tswj hwm hauv qab ntawm cov lus teb ntawm tes.
●Rigorous Quality Control: Peb siv cov ntsiab lus tseem ceeb hauv txhua theem, los ntawm cov qauv kev npaj mus rau kev npaj tsev qiv ntawv, kev sib txuas lus thiab bioinformatics. Peb cov kev soj ntsuam xyuas kom zoo xyuas kom cov khoom xa tuaj raws li cov txiaj ntsig zoo.
●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 rau hauv cov txheej txheem ntawm tes thiab cov txheej txheem molecular hauv qab cov lus teb, ua kom koj tau txais tag nrho cov ntaub ntawv muaj peev xwm hais txog koj qhov kev sim cov ntaub ntawv.
●Kev them nyiaj yug tom qab muag: Peb nkag siab tias yuav tsum muaj qhov tseem ceeb npaum li cas, yog vim li cas peb txoj kev cog lus txuas ntxiv mus 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 | Platform | Cov ntaub ntawv pom zoo | Cov ntaub ntawv QC |
| rRNA depleted directional tsev qiv ntawv | Illumina PE 150 | 10-16 Gb | Q30≥85% |
Nucleotides:
| Conc. (ng / μl) | Tus nqi (μg) | Purity | Kev ncaj ncees |
| ≥80 | ≥ 0.8 | OD 260/280 = 1.7-2.5 OD 260/230 = 0.5-2.5 Txwv tsis pub lossis tsis muaj protein ntau lossis DNA kis tau pom ntawm gel. | RIN ≥6.0; 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 lossis txoj hnyuv: 450 mg
Viscera lossis hlwb: 240 mg
Cov leeg: 600 mg
Pob txha, plaub hau lossis tawv nqaij: 1.5g
● Arthropods:
piv: 9g
Crustacea: 450 mg
● Cov ntshav tag nrho:2 cev
● Cells: 106 hlwb
● Ntshav thiab Plasmaib: 6ml
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
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
- Cov ntaub ntawv nyoos
- Cov ntaub ntawv QC
- Kev sib xyaw ntawm genome
- Gene qauv (Lwm txoj kev splicing, gene qauv optimization thiab tshiab noob kwv yees)
- Gene expression quantification
- Kev txheeb xyuas qhov sib txawv
- DEG annotation thiab enrichment + Differential qhia lncRNA hom noob noob
- Daim ntawv pov thawj
- lncRNA kev txheeb xyuas (lncRNA kev txuag thiab paub lncRNA)
- lncRNA lub hom phiaj kev kwv yees cov noob
- lncRNA kev qhia quantification
- Kev sib koom ua ke nrog mRNA cov ntaub ntawv
Differential Gene Expression (DEGs) tsom xam
Quantification of lncRNA expression – clustering
Enrichment ntawm lncRNA lub hom phiaj noob
Sib koom mRNA thiab lncRNA txoj hauj lwm tsom xam - Circos zajlus (lub voj voog nruab nrab yog mRNA thiab sab hauv cicrlce yog lncRNA)
Tshawb nrhiav cov kev nce qib uas tau pab txhawb los ntawm BMKGene's lncRNA equencing cov kev pabcuam los ntawm kev sau cov ntawv tshaj tawm.
Ji, H. et al. (2020) 'Kev txheeb xyuas, kev ua haujlwm kwv yees, thiab qhov tseem ceeb lncRNA pov thawj ntawm kev ntxhov siab ntsig txog lncRNAs hauv nas lub siab',Kev Tshaj Tawm Txog Kev Tshawb Fawb2020 10:1, 10(1), pp. 1–14. doi: 10.1038/s41598-020-57451-7 ib.
Jia, Z. et al. (2021) 'Integrative Transcriptomic Analysis Qhia Txog Kev Tiv Thaiv Kab Mob rau CyHV-3-Resistant Common Carp Strain',Frontiers hauv Immunology, 12, ib. 687151. doi: 10.3389/FIMMU.2021.687151/BIBTEX.
Wang, XJ et al. (2022) 'Multi-Omics Integration-Based Prioritization of Competing Endogenous RNA Regulation Networks in Small Cell Lung Cancer: Molecular Characteristics and Drug Candidates',Frontiers hauv Oncology, 12, ib. 904865. doi: 10.3389/FONC.2022.904865/BIBTEX.
Xiao, L. et al. (2020) 'Cov noob caj noob ces ntawm cov noob caj noob ces coexpression network hauv qab photosynthesis hauv Populus',Plant Biotechnology Journal, 18(4), pp. 1015–1026. doi: 10.1111/PBI.13270 ib.
Zheng H., et al. (2022) 'Lub Ntiaj Teb Kev Tswj Xyuas Network rau Dysregulated Gene Expression thiab Abnormal Metabolic Signaling in Immune Cells in Microenvironment of Graves' Disease thiab Hashimoto's Thyroiditis',Frontiers hauv Immunology, 13, ib. 879824. doi: 10.3389/FIMMU.2022.879824/BIBTEX.
