I-TRANSCRIPTOMICS
imvelo
EZOKUXHUMANA
Ukuchazwa okulotshiweyo okugcwele kokuguqulwa kwe-SF3B1 ku-chronic lymphocytic leukemia kuveza ukwehliswa kokulawulwa kwama-intron agciniwe
Imibhalo yobude obugcwele| Ukulandelana kwe-Nanopore| Ukuhlaziywa kwe-isoform okuhlukile
Ingemuva
Sukuguqulwa kwe-omatic ku-splicing factor SF3B1 kuye kwabikwa kabanzi ukuthi kuhlotshaniswa nomdlavuza ohlukahlukene, okuhlanganisa i-lymphocytic leukemia (CLL), uveal melanoma, umdlavuza webele, njll. Ngaphezu kwalokho, izifundo ze-transcriptomic ezifundeka kafushane ziye zembula amaphethini e-aberrant splicing abangelwa ukuguqulwa kwe-SF3B1. Kodwa-ke, izifundo zalawa maphethini okuhlanganisa ahlukile sekuyisikhathi eside zilinganiselwe ezingeni lomcimbi kanye nokuntuleka kolwazi ngeleveli ye-isoform ngenxa yomkhawulo wemibhalo ehlanganisiwe efundiwe kafushane. Lapha, iplathifomu yokulandelana kwe-nanopore yethulwa ukuze kukhiqizwe imibhalo yobude obugcwele, enike amandla ukuguqulwa kwama-isoform e-AS.
Idizayini Yokuhlola
Izivivinyo
Ukuhlanganisa:1. CLL-SF3B1(WT) 2. CLL-SF3B1(K700E ukuguqulwa); 3. Ama-B-cell ajwayelekile
Isu lokulandelanisa:Ukulandelana kwelabhulali ye-Minion 2D, ukulandelana komtapo wezincwadi we-PromethION 1D; idatha yokufunda okufushane evela kumasampuli afanayo
Inkundla yokulandelanisa:I-ONT MinION; I-ONT PromethION;
Ukuhlaziywa kwe-Bioinformatics
Imiphumela
Aisamba sezigidi ezingama-257 ezifundiwe zakhiwe kumasampula e-CLL angu-6 nama-B-cell angu-3. Ngokwesilinganiso esingu-30.5% salokhu kufundiwe kukhonjwe njengokulotshiweyo okugcwele ubude.
FUkuhlaziywa kwe-isoform ye-ull-length ye-RNA(FLAIR) kwathuthukiswa ukuze kukhiqizwe isethi yama-isoform okuzethemba okukhulu. I-FLAIR ingafingqwa kanje:
Nukuqondanisa okufundekayo kwe-anopore: thola ukwakheka okulotshiwe okujwayelekile okusekelwe ku-genome eyireferensi;
Sukulungiswa kwe-plice junction: amaphutha alungile okulandelana (okubomvu) nesayithi le-splice kusuka kuma-intrones anezichasiselo, izingeniso ezivela kudatha efundeka kafushane noma kokubili;
Ci-ollapse: fingqa ama-isoform amele asekelwe kumaketango okuhlangana kwe-splice(isethi yokudlula yokuqala). Khetha i-isofrom enokuzithemba okukhulu ngokusekelwe enanini lokufundwa okusekelayo(I-Threshold: 3).
Umfanekiso 1. Ukuhlaziywa kwe-FLAIR ukuze kutholakale ama-isoform e-lenth egcwele ahlotshaniswa nokuguqulwa kwe-SF3B1 ku-CLL
FI-LAIR Ihlonze ama-isoform ahlukene anokuzithemba angama-326,699, ama-90% awo angama-isoform anoveli. Iningi lalawa ma-isoform angachazwanga atholwe eyinhlanganisela yamanoveli ama-splice juntions (142,971), kanti amanye ama-isoforms anoveli aqukethe noma i-intron egciniwe (21,700) noma inoveli exon (3594).
Lukulandelana kokufunda isikhathi eside kunika amandla ukukhonjwa kwezindawo eziguquguqukayo ze-SF3B1-K700E -ezishintshiwe ezingeni le-isoform. Amanye ama-3'SS angu-35 kanye nama-5'SS angu-10 atholakala ehlukaniswe ngokuphawulekayo phakathi kwe-SF3B1-K700E kanye ne-SF3B1-WT. Okungu-33 kwezingu-35 okushintshiwe kusanda kutholwa ngokulandelana okufundwe isikhathi eside. Kudatha ye-Nanopore, ukusatshalaliswa kwebanga phakathi kwe-SF3B1-K700E-eshintshiwe engu-3'SS kuya eziqongweni ze-canonical sites cishe -20 bp, ehluke kakhulu ekusabalaliseni kokulawula, okufana nalokho okuye kwabikwa ngokulandelana kwe-CLL yokufunda okufushane. Ama-Isoform ofuzo lwe-ERGIC3 ahlaziywa, lapho i-isoform yenoveli equkethe isiza se-splice esisondelene yatholwa inala kakhulu ku-SF3B1-K700E. Kokubili i-proximal ne-distal 3'SS ibihlotshaniswa namaphethini e-AS ahlukanisiwe akhiqiza ama-isoform amaningi.
Umfanekiso 2. Amanye amaphethini okuhlanganisa angu-3′ akhonjwe nedatha yokulandelana kwe-nanopore
Ukuhlaziywa kokusetshenziswa komcimbi we-IR bekukhawulelwe isikhathi eside ekuhlaziyeni okusekelwe ekufundeni okufushane ngenxa yokuqiniseka ekuhlonzweni kwe-IR nokulinganisa. Ukuvezwa kwama-isoforms e-IR ku-SF3B1-K700E naku-SF3B1-WT alinganiswa ngokusekelwe ekulandeleni kwe-nanopore, okuveza ukulawulwa phansi komhlaba wonke kwama-isoforms e-IR ku-SF3B1-K700E.
Umfanekiso 4. Ukuqina kwezolimo kanye nokuxhumana kwenethiwekhi kuzo zonke izinhlelo zokulima ezintathu (A no-B); Ukuhlaziywa kwehlathi okungahleliwe(C) kanye nobudlelwano phakathi kokuqina kwezolimo kanye nekoloni ye-AMF (D)
Umfanekiso 3. Imicimbi yokugcinwa kwe-Intron yehliswe kakhulu ku-CLL SF3B1-K700E
Ubuchwepheshe
I-Nanopore Long-read Sequencing
Nukulandelana kwe-anopore kuwubuchwepheshe bokulandelanisa isignali kagesi yesikhathi sangempela se-molecule.
DI-DNA ene-ouble-stranded noma i-RNA izobophezela kuphrotheni e-nanoporous eshumekwe ku-biofilm futhi ivuleke ngaphansi kokuhola kwephrotheni yemoto.
DIzintambo ze-NA/RNA zidlula kuphrotheni yesiteshi se-nanopore ngenani elithile ngaphansi kwesenzo somehluko kagesi.
Mama-olecule akhiqiza amasignali kagesi ahlukene ngokuya ngesakhiwo samakhemikhali.
Rukutholwa kwesikhathi sangempela kokulandelana kufinyelelwa ngokushaya ucingo.
Ukusebenza kokulandelana kwe-transcriptome yobude obugcwele
√ Ukugcwaliswa Kwedatha
Ukufundwa okuphindwe izikhathi ezingu-7 okumbalwa okudingekayo ukuze kufinyelelwe ekugcwaliseni idatha okufanayo.
√ Ukuhlonza Isakhiwo Sombhalo
Ukukhonjwa kwezinhlobonhlobo zesakhiwo esivumelanayo sobude obugcwele bokufunda okulotshiwe ngakunye
√ Ukuhlaziya okuhlukile kwezinga lokulotshiweyo -Veza izinguquko ezifihlwe ukufundwa okufushane
Ireferensi
Tang AD , Soulette CM , Baren MJV , et al. Ukulingiswa okulotshiweyo okugcwele ubude be-SF3B1 kuguquko ku-lymphocytic leukemia engapheli kuveza ukwehliswa kwama-intron agciniwe[J]. Ukuxhumana Kwemvelo.
Ubuchwepheshe Nokugqama ihlose ukwabelana ngohlelo lokusebenza oluyimpumelelo lwakamuva kakhulu lobuchwepheshe bokulandelana komphumela ophezulu ezinkundleni ezihlukahlukene zokucwaninga kanye nemibono emihle ekuklanyweni kokuhlola kanye nokumbiwa kwedatha.
Isikhathi sokuthumela: Jan-08-2022