Raupapa mRNA Roa-Katoa-Nanopore
Ngā Āhuatanga
● Te hopu i te mRNA poly-A, ā, muri iho ka hangaia te cDNA me te whakarite whare pukapuka
● Te raupapatanga o ngā tuhinga roa katoa
● Tātaritanga koiora-pūtaiao i runga i te whakahāngaitanga ki tētahi ira tohutoro
● Ko te tātaritanga koiora-pūtaiao ehara i te mea ko te whakaaturanga i te taumata ira me te isoform anake engari ko te tātaritanga o te lncRNA, ngā whakakotahitanga ira, te poly-adenylation me te hanganga ira
Ngā Painga Ratonga
●Te ine i te whakapuakitanga i te taumata isoform: te whakahohe i te tātaritanga taipitopito me te tika o te whakapuakitanga, te hura i ngā huringa ka hunaia pea i te tātaritanga o te whakapuakitanga ira katoa
●Ngā Hiahia Raraunga kua Whakaitihia:Ki te whakatauritea ki te Raupapa Whakatipuranga-Panuku (NGS), he iti ake ngā whakaritenga raraunga mō te raupapatanga Nanopore, e āhei ai te whakatutuki i ngā taumata ōrite o te kīkī ira me ngā raraunga iti ake.
●Te tika ake o te ine i te whakapuakitangai te taumata ira me te taumata isoform
●Te tautuhi i ngā mōhiohio transcriptomic tāpiri: te polyadenylation rerekē, ngā ira whakakotahi me te lcnRNA me ō rātou ira ūnga
●Tohungatanga WhānuiHe nui ngā wheako o tā mātou tīma ki ia kaupapa, kua oti i a rātou neke atu i te 850 ngā kaupapa Nanopore transcriptome roa-katoa, ā, neke atu i te 8,000 ngā tauira i tukatukahia.
●Tautoko Whai muri i te Hokonga: e kore e mutu te kaupapa, engari e toru marama te roa o tā mātou whakapau kaha. I tēnei wā, ka tukuna e mātou he ratonga whai muri i te kaupapa, he āwhina rapurongoā, me ngā wā pātai me ngā whakautu hei whakautu i ngā pātai e pā ana ki ngā hua.
Ngā Whakaritenga Tauira me te Tukunga
| Whare Pukapuka | Rautaki raupapatanga | Ngā raraunga e taunakitia ana | Mana Kounga |
| Whakarei ake i te Poly A | Nanopore PromethION 48 | 6/12 GB | Kaute kounga toharite: Q10 |
Ngā Whakaritenga Tauira:
Ngā nukleotida:
| Te nui o te konupūmā (ng/μl) | Te nui (μg) | Te parakore | Ngākau pono |
| ≥ 100 | ≥ 1.0 | OD260/280=1.7-2.5 OD260/230=0.5-2.5 He iti noa iho, kāore rānei he poke o te pūmua, o te DNA rānei e kitea ana i runga i te tiēre. | Mō ngā tipu: RIN≥7.0; Mō ngā kararehe: RIN≥7.5; 5.0≥28S/18S≥1.0; te pikinga turanga iti, kāore rānei |
● Ngā tipu:
Pūtake, Kākano, Putiputi rānei: 450 mg
Rau, Kākano rānei: 300 mg
Hua: 1.2 karamu
● Kararehe:
Ngakau, Puku rānei: 300 mg
Ngā whekau, te roro rānei: 240 mg
Uaua: 450 mg
Ngā wheua, ngā makawe, te kiri rānei: 1 karamu
● Ngā Angaanga:
Ngā pepeke: 6g
Ngā Kōura: 300 mg
● Toto katoa1 ngongo
● Ngā pūtau106 ngā pūtau
Tukunga Tauira e Manakohia ana
Ipu: ngongo whakapūkara 2 ml (Kāore i te tūtohutia te pepa konumohe)
Tauira tapanga: Rōpū+tāruatia hei tauira A1, A2, A3; B1, B2, B3.
Tukunga:
1. Hukapapa maroke: Me tākai ngā tauira ki roto i ngā putea ka tanumia ki roto i te hukapapa maroke.
2. Ngā ngongo RNA-pūmau: Ka taea te whakamaroke i ngā tauira RNA i roto i te ngongo RNA-pūmau (hei tauira, RNA-pūmau®) ka tukuna i te pāmahana rūma.
Rere Mahi Ratonga
Ngā nukleotida:
Tukunga tauira
Te hanganga whare pukapuka
Te raupapatanga
Tātari raraunga
Ngā ratonga i muri i te hoko
Rere Mahi Ratonga
kiko:
Te hoahoa whakamātautau
Tukunga tauira
Te tango RNA
Te hanganga whare pukapuka
Te raupapatanga
Tātari raraunga
Ngā ratonga i muri i te hoko
● Te tukatuka raraunga mata
● Te tautuhi tuhinga
● Te honohono rerekē
● Te ine i te whakapuakanga i roto i te taumata ira me te taumata isoform
● Tātaritanga o te rerekētanga o te whakaaturanga
● Te tuhipoka me te whakarei ake i te mahi (ngā DEG me ngā DET)
Tātaritanga honohono rerekē
Tātaritanga Polyadenylation Rerekē (APA)
Matapae lncRNA
He whakamārama mō ngā ira hou
Te whakarōpūtanga o ngā DET
Ngā Whatunga Pūmua-Pūmua i roto i ngā DEG
Tūhurahia ngā whanaketanga e āwhinahia ana e ngā ratonga raupapatanga mRNA roa-katoa a Nanopore a BMKGene mā roto i tētahi kohinga pukapuka kua whiriwhiria.
Gong, B. et al. (2023) 'Te whakahohenga epigenetic me te transcription o te secretory kinase FAM20C hei oncogene i roto i te glioma', Journal of Genetics and Genomics, 50(6), pp. 422–433. doi: 10.1016/J.JGG.2023.01.008.
He, Z. et al. (2023) 'Ko te raupapatanga transcriptome roa-katoa o ngā lymphocytes e whakautu ana ki te IFN-γ e whakaatu ana i te urupare ārai mate Th1-skewed i roto i te pāpaka (Paralichthys olivaceus)', Fish & Shellfish Immunology, 134, wh. 108636. doi: 10.1016/J.FSI.2023.108636.
Ma, Y. et al. (2023) 'Te tātaritanga whakatairite o ngā tikanga raupapatanga RNA PacBio me ONT mō te tautuhi i te paitini Nemopilema Nomurai', Genomics, 115(6), wh. 110709. doi: 10.1016/J.YGENO.2023.110709.
Yu, D. et al. (2023) 'E whakaatu ana te tātaritanga nano-seq i te rerekētanga o te mahi i waenga i ngā exosomes me ngā microvesicles i ahu mai i te hUMSC', Stem Cell Research and Therapy, 14(1), pp. 1–13. doi: 10.1186/S13287-023-03491-5/TABLES/6.
