Uburebure bwuzuye mRNA ikurikirana-Nanopore
Inyungu za serivisi
ØKubogama gukabije
ØKugaragaza uburebure bwuzuye cDNA
ØAmakuru make asabwa kugirango yandike umubare winyandiko-mvugo
ØKumenyekanisha isoforms nyinshi kuri gen
ØKugereranya kugereranya murwego rwa isoform
Ibisobanuro bya serivisi
| Isomero | Ihuriro | Basabwe gutanga umusaruro (Gb) | Kugenzura ubuziranenge |
| cDNA-PCR (Poly-A ikungahaye) | Nanopore Isezerano P48 | 4 Gb / icyitegererezo (Ukurikije ubwoko) | Ikigereranyo cyuzuye ＞ 70% Ugereranyije amanota meza: Q10
|
Isesengura rya bioinformatics
ü Gutunganya amakuru mabi
üKumenyekanisha inyandiko
üUbundi gutera
üKugaragaza ingano murwego rwa gene na urwego rwa isoform
üIsesengura ry'imvugo itandukanye
üImikorere yo gutangaza no gukungahaza (DEGs na DETs)
Icyitegererezo gisabwa no gutanga
Icyitegererezo gisabwa:
Nucleotide:
| Isuku | Ubunyangamugayo | Umubare |
| OD260 / 280≥1.8 ； OD260 / 230≥1.0 peak Impinga nziza kuri 260 nmLimited cyangwa nta proteine cyangwa ADN yanduye yerekanwe kuri gel. | Kubimera: RIN≥7.5; Ku nyamaswa: RIN≥8; 28S / 18S≥1.0; | Umwanzuro.≥40 ng / μl; Igiteranyo ≥ 1 μg |
Tissue: Uburemere (bwumye): ≥1 g
* Kuri tissue ntoya ya mg 5, turasaba kohereza flash ya flash ikonje (muri azote yuzuye).
Guhagarika ingirabuzimafatizo: Kubara Akagari = 3 × 106- 1 × 107
* Turasaba kohereza lysate ya selile ikonje.Mugihe iyo selile ibara ntoya 5 × 105, flash yahagaritswe muri azote yuzuye, birasabwa gukuramo mikoro.
Ingero zamaraso: Volume≥1 ml
Basabwe Gutanga Icyitegererezo
Ibikoresho: ml 2 ya centrifuge (Tin foil ntabwo isabwa)
Icyitegererezo cyerekana: Itsinda + kwigana urugero A1, A2, A3;B1, B2, B3 ... ...
Kohereza: 1. Urubura-urubura: Ingero zigomba gupakirwa mumifuka hanyuma zigashyingurwa-urubura.
- Imiyoboro ya RNA: Ingero za RNA zirashobora gukama mumashanyarazi ya RNA (urugero RNAstable®) hanyuma ikoherezwa mubushyuhe bwicyumba.
Akazi ka serivisi
Icyitegererezo
Kubaka isomero
Urukurikirane
Isesengura ryamakuru
Serivisi nyuma yo kugurisha
Akazi ka serivisi
Igishushanyo mbonera
Icyitegererezo
Gukuramo RNA
Kubaka isomero
Urukurikirane
Isesengura ryamakuru
Serivisi nyuma yo kugurisha
1.Isesengura ry'imvugo itandukanye -Umugambi wibirunga
Isesengura ryerekana imvugo itandukanye irashobora gutunganywa murwego rwa gene kugirango tumenye genes zitandukanye (DEGs) no murwego rwa isoform kugirango tumenye bitandukanye.
inyandiko-mvugo yerekanwe (DETs)
2.Ubushuhe bukurikirana
3.Ikimenyetso cyo gutondeka no gutondeka
Ubwoko butanu bwubundi buryo bwo gutondeka ibintu bishobora guhanurwa na Astalavista.
4.Ubundi poly-adenylation (APA) ibyabaye biranga na Motif kuri 50 bp hejuru ya poly-A
Urubanza rwa BMK
Ubundi buryo bwo gutondeka hamwe na isoform-urwego rwo kugereranya na nanopore yuzuye-transcriptome ikurikiranye
Byatangajwe:Itumanaho rya Kamere, 2020
Ingamba zikurikirana:
Itsinda: 1. CLL-SF3B1 (WT);2. CLL-SF3B1 (K700E ihinduka);3. B-selile isanzwe
Ingamba zikurikirana: Minion 2D ikurikirana isomero, PromethION 1D ikurikirana isomero;bigufi-soma amakuru kuva kurugero rumwe
Urubuga rukurikirana: Nanopore MinION;Nanopore Gusezerana;
Ibisubizo by'ingenzi
1.Isoform-urwego Ubundi Gutandukanya Kumenyekanisha
Urutonde rwasomwe igihe kirekire rutanga imbaraga zo kumenya mutant SF3B1K700E-guhindura ibice byimbuga kuri isoform-urwego.35 ubundi buryo 3′SS na 10 ubundi 5′SS wasangaga bitandukanijwe cyane hagati ya SF3B1K700Ena SF3B1WT.33 kuri 35 byahinduwe byavumbuwe nuburyo bukurikiranye gusoma.
2.Isoform-urwego Ubundi Gutandukanya Kugereranya
Kugaragaza kugumana intron (IR) isoforms muri SF3B1K700Ena SF3B1WTbyapimwe hashingiwe kuri nanopore ikurikirana, byerekana isi yose munsi-kugenzura IR isoforms muri SF3B1K700E.
Reba
Tang AD, Soulette CM, Baren MJV, nibindi.Inyandiko-mvugo yuzuye iranga ihinduka rya SF3B1 muri lymphocytike ya leukemiya idakira igaragaza kugabanuka kwimitsi yagumishijwe [J].Itumanaho rya Kamere.
