Ukulandelelana kwe-mRNA okusekelwe ku-Non-Reference
Izici
● Ukuthwebula i-poly mRNA ngaphambi kokulungiselela umtapo wolwazi
● Kungakhathalekile ukuthi iyiphi i-genome ebhekisela kuyo: kusekelwe ekuhlanganisweni kwe-de novo kwemibhalo eqoshiwe, okudala uhlu lwama-unigenes abhalwe ngezincazelo zedatha eziningi (NR, Swiss-Prot, COG, KOG, eggNOG, Pfam, GO, KEGG)
● Ukuhlaziywa okuphelele kwe-bioinformatic kokubonakaliswa kwezakhi zofuzo kanye nesakhiwo sombhalo
Izinzuzo Zesevisi
●Ubuchwepheshe Obubanzi: ngomlando wokucubungula amasampula angaphezu kuka-600,000 kwa-BMKGENE, ahlanganisa izinhlobo ezahlukene zamasampula njengezakhi zamaseli, izicubu, kanye noketshezi lomzimba, ithimba lethu liletha ulwazi oluningi kuyo yonke iphrojekthi. Siqede ngempumelelo amaphrojekthi e-mRNA-Seq angaphezu kuka-100,000 emikhakheni eyahlukene yocwaningo.
●Ukulawulwa Kwekhwalithi Okuqinile: sisebenzisa amaphuzu okulawula ayisisekelo kuzo zonke izigaba, kusukela ekulungiseleleni amasampula kanye nomtapo wolwazi kuya ekulandeleni nasekubhaleni ulwazi nge-bioinformatics. Lokhu kuqapha ngokucophelela kuqinisekisa ukulethwa kwemiphumela esezingeni eliphezulu njalo.
● Isichasiselo Esiphelele: sisebenzisa izizindalwazi eziningi ukuze sichaze ngokusebenzayo i-Differentially Expressed Genes (DEGs) futhi senze ukuhlaziywa kokunotha okuhambisanayo, sinikeze ukuqonda ngezinqubo zamaseli kanye nama-molecule ezisekela impendulo ye-transcriptome.
●Usekelo Lwangemva Kokuthengisa: ukuzibophezela kwethu kudlulela ngale kokuqedwa kwephrojekthi ngesikhathi sesevisi yezinyanga ezintathu ngemuva kokuthengisa. Phakathi nalesi sikhathi, sinikeza ukulandelwa kwephrojekthi, usizo lokuxazulula izinkinga, kanye nezikhathi ze-Q&A ukuze kuxazululwe noma yimiphi imibuzo ehlobene nemiphumela.
Izidingo Zesampula Nokulethwa
| Umtapo Wolwazi | Isu lokulandelana kwezigaba | Kunconywa idatha | Ikhwalithi yokulawula |
| I-Poly A ecebile | I-Illumina PE150 I-DNBSEQ-T7 | 6-10 Gb | Q30≥85% |
Izidingo Zesampula:
Ama-Nucleotide:
| I-Conc.(ng/μl) | Inani (μg) | Ubumsulwa | Ubuqotho |
| ≥ 10 | ≥ 0.2 | OD260/280=1.7-2.5 OD260/230=0.5-2.5 Ukungcoliswa kwephrotheni noma i-DNA okulinganiselwe noma okungekho nhlobo kukhonjisiwe kujeli. | Kwezitshalo: I-RIN≥4.0; Ezilwaneni: I-RIN≥4.5; 5.0≥28S/18S≥1.0; ukuphakama okulinganiselwe noma okungekho kwesisekelo |
● Izitshalo:
Impande, Isiqu noma Ipetali: 450 mg
Iqabunga noma Imbewu: 300 mg
Izithelo: 1.2 g
● Isilwane:
Inhliziyo noma Amathumbu: 300 mg
I-Viscera noma Ubuchopho: 240 mg
Imisipha: 450 mg
Amathambo, Izinwele noma Isikhumba: 1g
● Ama-arthropod:
Izinambuzane: 6g
I-Crustacea: 300 mg
● Igazi eliphelele: Ishubhu eli-1
● Amaseli: 106 amaseli
Ukulethwa Kwesampula Okunconyiwe
Isitsha: Ishubhu le-centrifuge elingu-2 ml (Akukhuthazwa ukusebenzisa i-tin foil)
Ukulebula kwesampula: Iqembu + ukukopisha isib. A1, A2, A3; B1, B2, B3.
Ukuthunyelwa:
1. Iqhwa elomile: Amasampula kudingeka apakishwe ezikhwameni bese egqitshwa eqhweni elomile.
2. Amashubhu e-RNAstable: Amasampula e-RNA angomiswa eshubhu lokuzinzisa i-RNA (isb. i-RNAstable®) bese ethunyelwa ekushiseni kwegumbi.
Ukugeleza Komsebenzi Wesevisi
Umklamo wokuhlola
Ukulethwa kwesampula
Ukukhishwa kwe-RNA
Ukwakhiwa komtapo wolwazi
Ukulandelana
Ukuhlaziywa kwedatha
Izinsizakalo zangemuva kokuthengisa
I-Bioinformatics
Ukuhlanganiswa kwe-Transcriptome kanye nokukhethwa kwe-unigene
Isichasiselo se-Unigene
Ukuhambisana kwesampula kanye nokuhlolwa kokuphindaphindwa kwezinto eziphilayo
Izakhi Zofuzo Ezivezwe Ngokuhlukile (ama-DEG)
Isichasiselo Esisebenzayo sama-DEG
Ukuthuthuka Okusebenzayo kwama-DEG
Hlola intuthuko eyenziwe yizinsizakalo zokulandelana kwe-BMKGene ze-eukaryotic NGS mRNA ngokusebenzisa iqoqo lezincwadi ezikhethiwe.
UShen, F. et al. (2020) 'Ukuhlanganiswa kwe-transcriptome entsha kanye nokubonakaliswa kwezakhi zofuzo ezibandlulula ngokobulili kuma-gonad e-Amur catfish (Silurus asotus)', i-Genomics, 112(3), amakhasi 2603–2614. doi: 10.1016/J.YGENO.2020.01.026.
UZhang, C. et al. (2016) 'Ukuhlaziywa kokubhalwa phansi kwe-metabolism ye-sucrose ngesikhathi sokuvuvukala kwe-bulb kanye nokukhula ku-anyanisi (i-Allium cepa L.)', i-Frontiers in Plant Science, 7(Septhemba), ikhasi 212763. doi: 10.3389/FPLS.2016.01425/BIBTEX.
UZhu, C. nabanye. (2017) 'Ukuhlanganiswa okusha, ukuchazwa kanye nesichasiselo se-transcriptome ye-Sarcocheilichthys sinensis', PLoS ONE, 12(2). doi: 10.1371/JOURNAL.PONE.0171966.
UZou, L. nabanye. (2021) 'Ukuhlaziywa kwe-De novo transcriptome kunikeza ukuqonda ngokubekezelela usawoti kwe-Podocarpus macrophyllus ngaphansi kokucindezeleka kosawoti', i-BMC Plant Biology, 21(1), amakhasi 1–17. doi: 10.1186/S12870-021-03274-1/FIGURES/9.
