Sequencing ea Exome ea Motho e Felletseng
Likarolo tsa Tšebeletso
● Liphanele tse peli tsa exome tse fumanehang ho ipapisitse le ntlafatso ea sepheo ka li-probes: Sure Khetha Human All Exon v6 (Agilent) le xGen Exom Hybridization Panel v2 (IDT).
● Sequencing ho Illumina NovaSeq.
● Pipeline ea bioinformatiki e lebisitsoeng ho hlahloba mafu kapa ho hlahloba hlahala.
Melemo ea Tšebeletso
●Liphoso tsa Protein Coding Region: Ka ho hapa le ho hlophisa libaka tsa likhoutu tsa protheine, hWES e sebelisoa ho hlahisa mefuta e fapaneng e amanang le sebopeho sa protheine.
●Litšenyehelo tse Atlehang:hWES e fana ka hoo e ka bang 85% ea liphetoho tse amanang le lefu la motho ho tsoa ho 1% ea lefutso la motho.
●Ho Nepaha ha Phahameng: Ka botebo bo phahameng ba tatelano, hWES e thusa ho lemoha mefuta e fapaneng e tloaelehileng le mefuta e sa tloaelehang e nang le maqhubu a tlase ho 1%.
●Taolo ea Boleng e Tiileng: Re kenya tšebetsong lintlha tsa mantlha tsa taolo ho mekhahlelo eohle, ho tloha ho lisampole le boitokiso ba laeborari ho isa ho tatellano le bioinformatics. Tlhokomelo ena e hlokolosi e tiisa hore ho fanoa ka liphetho tsa boleng bo holimo kamehla.
●Tlhahlobo e Felletseng ea Bioinformatics: phaephe ea rona e fetela ka nģ'ane ho ho khetholla mefuta e sa tšoaneng ho genome ea litšupiso, kaha e kenyelletsa tlhahlobo e tsoetseng pele e etselitsoeng ho rarolla ka ho khetheha lipotso tsa lipatlisiso tse amanang le likarolo tsa lefutso tsa mafu kapa tlhahlobo ea tumor.
●Tšehetso ea Ka mor'a Thekiso:Boitlamo ba rona bo fetela ka nģ'ane ho ho phethoa ha morero ka nako ea tšebeletso ea likhoeli tse 3 ka mor'a thekiso. Nakong ena, re fana ka tatellano ea projeke, thuso ea ho rarolla mathata, le linako tsa Q&A ho araba lipotso life kapa life tse amanang le sephetho.
Litlhaloso tsa Mohlala
| Leano la Exon Capture | Leano la Tatelano | Tlhahiso ea data e khothalelitsoeng |
| Sure Select Human All Exon v6 (Agilent) kapa xGen Exom Hybridization Panel v2 (IDT)
| Illumina NovaSeq PE150 | 5 -10 Gb Bakeng sa mathata a mende/mafu a sa tloaelehang:> 50x Bakeng sa mehlala ea hlahala: ≥ 100x |
Litlhoko tsa Mohlala
| Mofuta oa Mohlala
| Chelete(Qubit®)
| Ho tsepamisa maikutlo | Bolumo
| Bohloeki(NanoDrop™) |
|
Genomic DNA
| ≥ 50 ng | ≥ 6 ng/μL | ≥ 15 μL | OD260/280=1.8-2.0 ha ho nyenyefatso, ha ho tšilafalo
|
Bioinformatics
Tlhahlobo ea bioinformatiki ea lisampole tsa lefu la hWES e kenyelletsa:
● Ho latela data QC
● Reference Genome Alignment
● Ho khetholla li-SNP le li-InDels
● Tlhaloso e sebetsang ea SNPs le InDels
Tlhahlobo ea bioinformatics ea lisampole tsa tumor e kenyelletsa:
● Ho latela data QC
● Reference Genome Alignment
● Ho khetholla li-SNP, li-InDels le mefuta e sa tšoaneng ea somatic
● Ho khetholla mefuta ea likokoana-hloko
● Ho hlahlojoa ha liphatsa tsa lefutso
● Ho tsebahatsa liphatsa tsa lefutso tse itšetlehileng ka liphetoho tse sebetsang hantle
● Tlhaloso ea phetoho boemong ba ho ba kotsing ea lithethefatsi
● Heterogeneity analysis - palo ea bohloeki le ploidy
Phallo ea Mosebetsi oa Ts'ebeletso
Mohlala oa thomello
Ho ntšoa ha DNA
Kaho ea laebrari
Tatelano
Tlhahlobo ea lintlha
Phano ea data
Litšebeletso tsa ka mor'a thekiso
QC ea data - Lipalopalo tsa ho ts'oaroa ha Exome
Boitsebiso bo fapaneng - InDels
Tlhahlobo e tsoetseng pele: ho tsebahatsa le ho ajoa ha li-SNP/InDels tse senyehileng - Circos plot
Tlhahlobo ea tumor: ho tsebahatsa le ho ajoa ha liphetoho tsa somatic - morero oa Circos
Tlhahlobo ea hlahala: mela ea clonal
