TS'ELISO
tlhaho
LITŠOANTŠISO
Sengoliloeng sa bolelele bo felletseng ba phetoho ea SF3B1 ho chronic lymphocytic leukemia e senola ho theola taolo ea li-intron tse bolokiloeng.
Lingoliloeng tsa bolelele bo felletseng|Nanopore sequencing|Tlhahlobo e 'ngoe ea isoform
Ka morao
Sliphetoho tsa omatic in splicing factor SF3B1 e 'nile ea tlalehoa ka bophara hore e amahanngoa le mefuta e sa tšoaneng ea kankere, ho akarelletsa le lefu le sa foleng la lymphocytic leukemia(CLL), uveal melanoma, kankere ea matsoele, joalo-joalo. Ho phaella moo, liphuputso tse khutšoanyane tse balang li-transcriptomic li senotse mekhoa e sa tšoaneng ea splicing e bakoang ke liphetoho tsa SF3B1.Leha ho le joalo, liphuputso tse mabapi le mekhoa ena e meng ea ho arola li 'nile tsa lekanyetsoa ho boemo ba liketsahalo le ho hloka tsebo ka boemo ba isoform ka lebaka la moeli oa litlaleho tse kopantsoeng tse baloang hakhutšoanyane.Mona, sethala sa tatellano ea nanopore se ile sa hlahisoa ho hlahisa lingoloa tse bolelele bo felletseng, tse ileng tsa matlafatsa ts'ebeliso ea li-isoform tsa AS.
Moqapi oa Teko
Liteko
Ho hlopha:1. CLL-SF3B1 (WT) 2. CLL-SF3B1 (K700E phetoho);3. Lisele tsa B tse tloaelehileng
Leano la tatellano:tatellano ea laeborari ea MinION 2D, tatellano ea laeborari ea PromethION 1D;lintlha tse khutšoane tse baloang ho tsoa lisampoleng tse tšoanang
Sequencing platform:ON MinION;ONT PromethION;
Tlhahlobo ea Bioinformatics
Liphetho
Akakaretso ea lipalo tse limilione tse 257 li entsoe ho tsoa ho lisampole tse 6 tsa CLL le lisele tse 3 tsa B.Ka karolelano ea 30.5% ea lipalo tsena li ile tsa khetholloa e le likopi tsa bolelele bo felletseng.
FUll-length alternative isoform analysis of RNA(FLAIR) e ile ea ntlafatsoa ho hlahisa sehlopha sa li-isoform tse nang le boitšepo bo phahameng.FLAIR e ka akaretsoa joalo ka:
Nanopore bala teka-tekano: hlwaya sebopeho sa sengoloa se akaretsang se ipapisitseng le genome ya referense;
Splice junction correction: liphoso tse nepahetseng tsa tatelano (tse khubelu) le sebaka sa splice ho tsoa ho li-introns tse hlalositsoeng, li-introne ho tsoa ho data e baloang hanyane kapa ka bobeli;
Collapse: akaretsa li-isoform tsa moemeli tse thehiloeng ho liketane tsa mateano a marang-rang (sete ea pele ea pele).Khetha isofrom e nang le ts'epo e kholo ho latela palo ea lipalo tse tšehetsang (Threshold: 3).
Setšoantšo sa 1. Tlhahlobo ea FLAIR ho khetholla li-isoform tse feletseng tse amanang le phetoho ea SF3B1 ho CLL
FLAIR e Khetholla li-isoform tse 326,699 tse nang le boitšepo bo phahameng, tseo 90% ea tsona e leng li-isoform tse ncha.Boholo ba li-isoform tsena tse sa hlalosoang li fumanoe e le motsoako o mocha oa li-splice juntions (142,971), athe li-isoform tse ling kaofela li ne li na le intron (21,700) kapa novel exon (3594).
Ltatellano ea nako e telele e fana ka matla ho tsebahatsa libaka tse fetotsoeng tsa SF3B1-K700E -altered splice sebakeng sa isoform-level.Li-3'SS tse 35 le tse 10 tse ling tsa 5'SS li fumanoe li arotsoe ka tsela e fapaneng haholo lipakeng tsa SF3B1-K700E le SF3B1-WT.Liphetoho tse 33 ho tse 35 li sa tsoa sibolloa ka tatellano e baloang nako e telele.Litabeng tsa Nanopore, kabo ea sebaka se pakeng tsa SF3B1-K700E-e fetotsoeng 3'SS ho litlhōrō tsa libaka tsa canonical e pota-potile -20 bp, e fapaneng haholo le kabo ea taolo, e ts'oanang le e tlalehiloeng ka tatellano e khutšoanyane ea CLL.Li-Isoforms tsa liphatsa tsa lefutso tsa ERGIC3 li ile tsa hlahlojoa, moo buka ea isoform e nang le sebaka sa proximal splice se fumanoeng se le ngata ho SF3B1-K700E.Ka bobeli proximal le distal 3'SS li ne li amahanngoa le mekhoa e ikhethileng ea AS e hlahisang li-isoform tse ngata.
Setšoantšo sa 2. Mefuta e meng ea 3′ ea ho kopanya e khetholloang ka tatellano ea data ea nanopore
Tlhahlobo ea ts'ebeliso ea ketsahalo ea IR esale e lekantsoe ka nako e telele tlhahlobong e baloang ka nako e khuts'oane ka lebaka la ts'epo mabapi le boitsebahatso ba IR le lipalo.Pontšo ea li-isoform tsa IR ho SF3B1-K700E le SF3B1-WT li ne li lekantsoe ho ipapisitsoe le tatellano ea nanopore, e senola taolo e tlase ea lefats'e ea li-isoform tsa IR ho SF3B1-K700E.
Setšoantšo sa 4. Matla a temo le khokahanyo ea marang-rang ho pholletsa le mekhoa e meraro ea temo (A le B);Tlhahlobo ea meru e sa reroang(C) le Kamano lipakeng tsa matla a temo le bokolone ba AMF (D)
Setšoantšo sa 3. Liketsahalo tsa ho boloka li-intron li fokotsehile ka matla ho CLL SF3B1-K700E
Thekenoloji
Nanopore Long-read Sequencing
Nanopore sequencing ke molek'hule e le 'ngoe ea nako ea sebele ea theknoloji ea tatellano ea matšoao a motlakase.
DDNA ea ouble-stranded kapa RNA e tla tlama ho protheine ea nanoporous e kentsoeng ka har'a biofilm ebe e phutholoha tlas'a ketapele ea protheine ea motlakase.
DLikhoele tsa NA/RNA li feta protheine ea kanal ea nanopore ka sekhahla se itseng tlasa ketso ea phapang ea voltage.
Mli-olecules li hlahisa lipontšo tse fapaneng tsa motlakase ho latela sebopeho sa lik'hemik'hale.
Rphumano ea nako le nako ea tatellano e fihlelleha ka mehala ea motheo.
Ts'ebetso ea tatellano ea bolelele bo felletseng ba transcriptome
√ Phatlalatso ya data
Ho baloa ho fokolang ha 7 ho hlokahala ho fihlela kakaretso ea data e ka bapisoang.
√ Boitsebiso ba Sebopeho sa Sengoloa
Khethollo ea mefuta e fapaneng ea libopeho le tumellano ea bolelele bo felletseng ba sengoloa ka seng
√ Tshekatsheko ya phapano ya boemo ba dingolwa -Bontsha diphetoho tse patilweng ke dipalo tse kgutshwane
Referense
Tang AD , Soulette CM , Baren MJV , et al.Tlhaloso ea bolelele bo felletseng ba phetoho ea SF3B1 ho chronic lymphocytic leukemia e senola ho theoha ha li-introns tse bolokiloeng[J].Tlhaho Puisano.
Theknoloji le Lintlha-khōlō e ikemiselitse ho arolelana ts'ebeliso ea morao-rao e atlehileng ea mekhoa e fapaneng ea tatellano ea maemo a holimo mabaleng a fapaneng a liphuputso hammoho le mehopolo e bohlale moralong oa liteko le ho rafa data.
Nako ea poso: Jan-08-2022