China Eukaryotic mRNA ukulandelana-Illumina ifektri nabakhiqizi |BMK

I-Eukaryotic mRNA ukulandelana-Illumina

Eukaryotic mRNA sequencing-Illumina Featured Image

Ukulandelana kwe-mRNA kunika amandla ukuphrofayela wawo wonke ama-mRNA alotshwe kusuka kumaseli ngaphansi kwezimo ezithile.Ubuchwephesha obunamandla bokuveza iphrofayili yesisho sofuzo, izakhiwo zofuzo nezindlela zamangqamuzana ezinqubo ezithile zebhayoloji.Kuze kube manje, ukulandelana kwe-mRNA sekusetshenziswe kabanzi ocwaningweni oluyisisekelo, ekuxilongweni komtholampilo, ekuthuthukisweni kwezidakamizwa, njll.

Ipulatifomu: Illumina NovaSeq 6000

Xhumana nathi

Imininingwane Yesevisi

I-Bioinformatics

Imiphumela yedemo

Izinzuzo

ØAbanolwazi Kakhulu: Amasampula angaphezu kuka-200,000 asetshenzwe ku-BMK ehlanganisa izinhlobo zamasampula ahlukahlukene, okuhlanganisa isiko lamaseli, izicubu, uketshezi lomzimba, njll. kanye namaphrojekthi angaphezu kuka-7,000 mRNA-Seq avaliwe ahlanganisa izindawo ezihlukahlukene zocwaningo.

ØIsistimu yokulawula ikhwalithi eqinile: Amaphuzu okulawula ikhwalithi abalulekile ngazo zonke izinyathelo ezihlanganisa ukulungiswa kwesampula, ukulungiswa komtapo wolwazi, ukulandelana kanye ne-bioinformatics angaphansi kokuqapha ukuze kulethwe imiphumela yekhwalithi ephezulu.

ØImininingo egciniwe eminingi etholakalayo yesichasiselo somsebenzi kanye nezifundo zokucebisa ukufeza izinjongo zocwaningo ezihlukahlukene.

ØAmasevisi angemuva kokuthengisa: Izinsizakalo ezithengiswa ngemva kokuthengisa zivumeleke izinyanga ezi-3 lapho iphrojekthi isiqediwe, okuhlanganisa ukulandelela amaphrojekthi, ukuxazulula izinkinga, imiphumela ye-Q&A, njll.

Izidingo Zesampula Nokulethwa

Umtapowolwazi	Isu lokulandelanisa	Idatha iyanconywa	Ikhwalithi yokulawula
I-Poly A inothisiwe	I-Ilumina PE150	6 gb	Q30≥85%

Izimfuneko zesampula:

Ama-Nucleotides:

Ubumsulwa	Ubuqotho	Inani
OD260/280≥1.7-2.5 OD260/230≥0.5-2.5Amaprotheni anomkhawulo noma awekho nhlobo noma amagciwane e-DNA aboniswa kujeli.	Okwezitshalo: RIN≥6.5;Ezilwaneni: RIN≥7;28S/18S≥1.0;ukuphakama okulinganiselwe noma okungekho	Conc.≥30 ng/μl;Ivolumu ≥ 10 μl;Isamba ≥ 1.5 μg

Izicubu: Isisindo(somile):≥1 g
*Ngezicubu ezincane kuno-5 mg, sincoma ukuthi uthumele isampula yezicubu eziqandisiwe (ku-nitrogen ewuketshezi).

Ukumiswa kweseli:Inani lamaseli = 3×10⁶- 1 × 10⁷
*Sincoma ukuthi uthumele i-frozen cell lysate.Esimeni lapho lelo seli liba lincane kuno-5×105, i-flash efriziwe ku-nitrogen ewuketshezi iyanconywa, okungcono kakhulu ekukhishweni okuncane.

Amasampula egazi:Umthamo≥1 ml

I-Microorganism:Isisindo ≥ 1 g

Ukulethwa Kwesampula Okunconyiwe

Isitsha: 2 ml centrifuge tube (Tin foil ayinconywa)

Isampula yokulebula: Ukuphindaphinda kweqembu+isb A1, A2, A3;B1, B2, B3... ...

Ukuthunyelwa:

Iqhwa elomile: Amasampula adinga ukupakishwa ezikhwameni futhi angcwatshwe eqhweni elomile.
Amashubhu e-RNAstable: Amasampula e-RNA angomiswa ngeshubhu lokuzinzisa le-RNA(isb. i-RNAstable®) futhi athunyelwe ngegumbi lokushisa.