ABANTU B'UMUNTU
imiterere y'ibinyabuzima
Urutonde rwasomwe igihe kirekire rugaragaza GGC gusubiramo kwaguka muri NOTCH2NLC ifitanye isano n'indwara ya neuronal intran nuclear
KUBONA ONT |Illumina |Byose bikurikirana |CRISPR-Cas9 ONT ikurikiranye |RNA-seq |ONT 5mC guhamagara methylation
Ingingo z'ingenzi
1.Koresheje isesengura rya Linkage kumuryango munini wa NIID, hagaragaye uturere tubiri duhujwe.
2.Kudashingira kumurongo usomye hamwe na Cas-9 byahujwe no gukungahaza ONT ikurikiranye yavumbuye impamvu ishobora kuba itera NIID, GGC gusubiramo kwaguka muri 5 ′ UTR ya NOTCH2NLC.Ubu bushakashatsi bwatangaje ko bwongeye kwaguka muri gen zihariye zabantu ku nshuro ya mbere byahindutse binyuze mu kwigana ibice.
3.Urutonde rwa RNA rwerekanye inyandiko zidasanzwe za antisense muntangiriro cyangwa imbere muri GGC gusubiramo uturere twagutse muri NOTCH2NLC.
Amavu n'amavuko
NIndwara ya euronal intranuclear (NIID) nindwara itera imbere kandi yica neurodegenerative, irangwa no kuba hariho eosinophilique hyaline intranuclear yinjira muri sisitemu yo hagati na peripheri.Kugaragara kwa clinique ihindagurika cyane itera ingorane zikomeye mugupima kugeza biopsy y'uruhu.Nyamara, uburyo bushingiye kuri histopathology buracyafite uburwayi bwo kwisuzumisha nabi, busaba ko hasobanurwa ubwoko bwa NIID.
Ibyagezweho
Isesengura ry'ihuza
SHort-soma ikurikiranye ishingiye kuri genome yose ikurikirana (WGS) hamwe na exome ikurikiranye (WES) yakorewe mumuryango munini wa NIID (13 bagize ingaruka nabanyamuryango 7 batagize ingaruka).Isesengura ryihuza kuri SNPs ryakuwe muri aya makuru ryerekanye uturere tubiri duhujwe gusa: akarere ka 3.5 Mb kuri 1p36.31-p36.22 (ntarengwa LOD = 2.32) nakarere ka 58.1 Mb kuri 1p22.1-q21.3 (LOD ntarengwa: 4.21 ).Nyamara, nta SNPs cyangwa CNV zitera indwara byagaragaye muri utwo turere duhujwe.
GGC gusubiramo kwaguka muri NOTCH2NLC
NUrutonde rushingiye kuri anopore rwatunganijwe ku 13 bagize ingaruka hamwe n’abanyamuryango 4 batagize ingaruka ku miryango 8 (undi munyamuryango wagize ingaruka yakurikiranwe na Pacbio igihe kirekire cyo gusoma.)Amakuru asomwe kuva kera yerekanaga indwara zifitanye isano na GGC gusubiramo kwaguka muri 5 ′ UTR ya NOTCH2NLC yerekana ikarita ya 58.1 Mb ihuza akarere (Ishusho 1).Uku kwaguka kwagaragaye no mubibazo 40 byose bya NIID byageragejwe na RP-PCR.
Cas-9 ihuza intego ikurikiranye kuri nanopore platform yakoreshejwe kugirango igere kumurongo wo gusoma kuri NOTCH2NLC gusubiramo (100 X-1,795 X).Urutonde rwumvikanyweho rwemeje neza nubushakashatsi bwibanze kuri GGC gusubiramo kwaguka.Byongeye kandi, {(GGA) n (GGC) n} gusubiramo byagaragajwe nkibimenyetso byerekana ubwoko bwa fenotipike yiganje (Ishusho 2).
Igicapo 1. Indwara zifitanye isano no kwaguka zagaragaye kuri exon 1 ya NOTCH2NLC isoforms.
Igishushanyo 2. Urutonde rwumvikanyweho rwa NPTCH2NLC rusubiramo abarwayi ba NIID bafite (*) cyangwa badafite intege nke-yiganje
NOTCH2NL ni genes yihariye yumuntu, ikekwa ko igira uruhare runini mubwihindurize bwubwonko bwabantu nindwara zifata ubwonko.Nyamara, genes eshatu zijyanye na NOTCH2 (NOTCH2NLA, NOTCH2NLB na NOTCH2NLC) zifite> 99.1% zikurikirana ntizakemuwe kugeza igihe inteko ya genome iheruka.Synthesis-yubusa kandi isomwa igihe kirekire kurubuga rwa nanopore yerekanye ibyiza bigaragara mugukemura uturere dusa cyane kandi (GGC) n gusubiramo hamwe na GC ikungahaye 100%.
GGC gusubiramo kwaguka muri NOTCH2NLC
TUrutonde rwa ranscriptome rwatunganijwe kubanyamuryango 2 bagizweho ingaruka nabanyamuryango 2 batagize ingaruka.Ubusanzwe gusoma byimbitse byabazwe kubisobanuro hamwe na antisense imirongo yo hejuru ya exons ya mbere ya NOTCH2NL.Inyandiko-mvugo idasanzwe yo kurwanya anti-sens yabonetse gusa mubibazo byibasiwe, bicara mugitangiriro cyangwa imbere mugusubiramo kwaguka (Impinga yumutuku muri F1-14 na F1-16 mumashusho 3.).Hiyongereyeho, DEGs 54 zamenyekanye kandi zose zarakungahajwe mu magambo ya GO na MPO bijyanye n'imikorere ya neuronal.
Igicapo 3. Ubusanzwe soma ubujyakuzimu hejuru yimbere ya exon ya mbere ya NOTCH2NLC mubitagize ingaruka (hejuru) kandi byagize ingaruka (munsi).
Ikoranabuhanga
Oxford Nanopore Teghnologies (ONT)
NUrutonde rwa anopore rwitandukanya nizindi mbuga zikurikirana, muburyo nucleotide isomwa neza nta sisitemu ya synthesis.Nkuko umugozi umwe ADN inyura muri pine ya nano nini (nanopore), nucleotide zitandukanye zitanga imiyoboro itandukanye ya ionic, ishobora gufatwa ikoherezwa muburyo bukurikirana.Ihuriro rya ONT ubwaryo ntirigaragaza imipaka ya tekinike ku burebure bwo gusoma ADN.Kubwibyo, Ultra-ndende isoma (ULRs) irahari kugirango genome ikorwe neza.Byongeye kandi, ibi bisomwa birebire cyane, birebire bihagije kugirango byambukiranya ibintu bikurikirana cyangwa imiterere itandukanye, bifasha gutsinda imbogamizi zo gusoma-bigufi hano.
Urutonde rwa Nanopore
Imiterere itandukanye (SV)
Synthesis-yubusa ikurikiranye ahanini yabitswe ADN methylation yamakuru kuri template.Methylated A, T, C na G ibyara ionic bigenda bitandukana na methylated, bishobora gusomwa neza na platifomu.Ikurikiranyabihe rya Nanopore riha imbaraga genome zose zerekana 5mC na 6mA kumurongo umwe wa nucleotide.
Reba
Jun Sone, n'abandi.al.Urutonde rwasomwe igihe kirekire rugaragaza GGC isubiramo kwaguka muri NOTCH2NLC ifitanye isano n'indwara ya neuronal intran nuclear.Kamere Kamere (2019)
Ikoranabuhanga hamwe nibyingenzi igamije gusangira ibyagezweho vuba muburyo butandukanye bwo gukoresha tekinoroji ikurikirana muburyo butandukanye hamwe nibitekerezo byiza mugushushanya no gucukura amakuru.
Igihe cyo kohereza: Mutarama-06-2022