Kev Tshawb Fawb Txog Qhov Tshwj Xeeb (Specific-Locus Amplified Fragment Sequencing) (SLAF-Seq)
Kev Ua Haujlwm
Cov kev pabcuam muaj qee qhov kev tsim ua ntej hauv silico los lav qhov kev xaiv enzyme zoo tshaj plaws ntawm kev npaj ntawm lub tsev qiv ntawv.
Txoj Kev Npaj Txuj Ci
Cov yam ntxwv ntawm kev pabcuam
● Kev txheeb xyuas kab mob ntawm NovaSeq nrog PE150.
● Kev npaj tsev qiv ntawv nrog ob chav barcoding, ua rau muaj ntau dua 1000 tus qauv sib sau ua ke.
● Ywj pheej ntawm cov genome siv:
Nrog rau kev siv genome: SNP thiab InDel kev tshawb pom
Tsis muaj cov genome siv: kev sib sau ua ke ntawm cov qauv thiab kev tshawb pom SNP
● Nyob rau hauvhauv-silicoua ntej tsim cov kev sib xyaw ua ke ntawm ntau yam enzyme txwv tsis pub dhau theem raug tshuaj xyuas kom pom cov uas tsim kom muaj kev faib tawm ntawm SLAF tags raws li genome.
● Thaum lub sijhawm ua ntej sim, peb qhov kev sib xyaw ua ke ntawm cov enzyme raug sim hauv 3 qhov qauv los tsim 9 lub tsev qiv ntawv SLAF, thiab cov ntaub ntawv no raug siv los xaiv qhov kev sib xyaw ua ke ntawm cov enzyme txwv tsis pub tshaj plaws rau qhov project.
Cov Kev Pabcuam Zoo
●Kev Tshawb Pom Cov Cim Qhia Txog Genetic SiabPeb siv lub tshuab barcode ob chav uas muaj peev xwm ua tau ntau yam kev ntsuas tib lub sijhawm, thiab kev ua kom lub tshuab ua haujlwm tau zoo dua qub, kom ntseeg tau tias cov lej cim npe ua tau raws li ntau yam kev xav tau ntawm ntau yam kev tshawb fawb.
● Kev vam khom tsawg rau ntawm GenomeNws tuaj yeem siv rau cov tsiaj txhu uas muaj lossis tsis muaj genome siv.
●Kev Tsim Qauv Yooj YimKev zom cov enzyme ib zaug xwb, ob chav enzyme, ntau hom enzyme, thiab ntau hom enzymes tuaj yeem xaiv tau los ua kom tau raws li ntau lub hom phiaj tshawb fawb lossis ntau hom tsiaj.
● Kev Ua Haujlwm Zoo Hauv Kev zom zaub mov Enzymatic: Kev sib txuas ntawm ib qhohauv-silicokev tsim ua ntej thiab kev sim ua ntej ua kom ntseeg tau tias kev tsim zoo tshaj plaws nrog kev faib tawm ntawm SLAF tags ntawm cov chromosome sib npaug (1 SLAF tag/4Kb) thiab txo qhov rov ua dua (<5%).
●Kev Paub Ntau YamPeb coj ntau yam kev paub dhau los rau txhua qhov project, nrog rau cov ntaub ntawv keeb kwm ntawm kev kaw ntau dua 5000 qhov project SLAF-Seq ntawm ntau pua hom tsiaj, suav nrog cov nroj tsuag, cov tsiaj txhu, cov noog, cov kab, thiab cov tsiaj txhu hauv dej.
● Kev Ua Haujlwm Bioinformatic uas Tus Kheej TsimPeb tau tsim ib txoj haujlwm bioinformatic workflow rau SLAF-Seq kom ntseeg tau tias qhov tso zis kawg muaj kev ntseeg siab thiab raug.
Cov Lus Qhia Txog Kev Pabcuam
| Hom kev tshuaj xyuas | Cov pejxeem pom zoo | Txoj kev npaj ua sequencing | |
| Qhov tob ntawm kev txheeb xyuas cov cim npe | Tus lej cim npe | ||
| Cov Ntawv Qhia Txog Genetic | 2 niam txiv thiab >150 tus menyuam | Niam Txiv: 20x WGS Kev Tawm Tsam: 10x | Qhov loj ntawm genome: <400 Mb: WGS raug pom zoo <1Gb: 100K daim ntawv cim npe 1-2Gb:: 200K cim npe >2Gb: 300K daim ntawv cim npe Cov cim npe siab tshaj 500k |
| Kev Tshawb Fawb Txog Kev Koom Tes Thoob Plaws Genome (GWAS) | ≥200 cov qauv | 10x | |
| Kev Hloov Pauv ntawm Genetic | ≥30 cov qauv, nrog rau >10 cov qauv los ntawm txhua pawg me | 10x | |
Cov Kev Cai rau Kev Pabcuam
Kev Ntsuas ≥ 5 ng/µL
Tag nrho cov nyiaj ≥ 80 ng
Agarose gel: tsis muaj lossis tsawg dua kev puas tsuaj lossis kev ua qias tuaj
Kev Xa Qauv Pom Zoo
Thawv: 2 ml centrifuge raj
(Rau feem ntau ntawm cov qauv, peb xav kom tsis txhob khaws cia rau hauv ethanol)
Kev Sau Npe Qauv: Cov qauv yuav tsum tau sau kom meej thiab zoo ib yam li daim ntawv qhia cov qauv uas tau xa tuaj.
Kev Xa Khoom: Dej khov qhuav: Cov qauv yuav tsum tau ntim rau hauv hnab ua ntej thiab faus rau hauv dej khov qhuav.
Kev Ua Haujlwm Pabcuam
Qauv QC
Kev sim tsav dav hlau
Kev sim SLAF
Kev Npaj Tsev Qiv Ntawv
Kev txheeb xyuas kab ke
Kev Tshawb Fawb Txog Cov Ntaub Ntawv
Kev Pab Tom Qab Muag Khoom
Peb qhov kev tshuaj xyuas bioinformatical muaj xws li:
Kev QC cov ntaub ntawv thiab kev txiav cov ntaub ntawv kom tshem tawm cov nyeem N-nplua nuj, cov nyeem adapter lossis cov nyeem tsis zoo.
Kev tswj xyuas qhov zoo thib ob ntawm qhov huv si nyeem los xyuas qhov kev faib tawm ntawm lub hauv paus, qhov zoo ntawm cov kab ke thiab kev ntsuam xyuas cov ntaub ntawv, tab sis kuj los xyuas qhov ua tau zoo ntawm kev zom zaub mov thiab cov ntawv ntxig tau txais.
Thaum cov ntawv nyeem tiav lawm, muaj ob txoj kev xaiv:
- Kev kos duab rau cov genome siv
- Tsis muaj ib qho genome siv: clustering
Tom qab nws, qhov kev tshuaj xyuas ntawm SLAF cov cim npe yog siv los ua qee qhov kev hu sib txawv los pab nrog kev tshawb pom cov cim: SNP, InDel, SNV, CV hu thiab annotation.
Kev faib tawm ntawm SLAF cim npe ntawm chromosomes:
Kev faib tawm ntawm SNPs ntawm cov chromosomes:
Cov lus piav qhia ntawm SNP
Jiang S, Li S, Luo J, Wang X thiab Shi C (2023) QTL mapping thiab transcriptome kev tshuaj xyuas ntawm cov suab thaj thaum lub sijhawm txiv hmab txiv ntoo siav ntawmPyrus pyrifolia.Pem Hauv Ntej. Kev Tshawb Fawb Txog Cog Qoob Loo.14: 1137104 ib. doi: 10.3389/fpls.2023.1137104 ib.
Li, J., Zhang, Y., Ma, R., Huang, W., Hou, J., Fang, C., & Sun, L. (2022). Kev txheeb xyuas ntawm st1 qhia txog kev xaiv uas cuam tshuam nrog kev caij tsheb ntawm cov noob morphology thiab cov roj thaum lub sijhawm cog taum pauv.Phau Ntawv Xov Xwm Txog Kev Siv Tshuaj Biotechnology, 20(6), 1110-1121. https://doi.org/10.1111/pbi.13791
Xu, P., Zhang, X., Wang, X.thiab lwm tus.Cov kab ke genome thiab kev sib txawv ntawm cov noob caj noob ces ntawm cov ntses carp,Cyprinus carpio.Nat Genet 46, 1212–1219 (2014). https://doi.org/10.1038/ng.3098
Zhuang, W., Chen, H., Yang, M.thiab lwm tus.Cov genome ntawm cov txiv laum huab xeeb uas cog rau hauv av muab kev nkag siab txog cov karyotypes ntawm cov noob taum, kev hloov pauv ntawm polyploid thiab kev yug tsiaj ntawm cov qoob loo.Nat Genet 51, 865–876 (2019). https://doi.org/10.1038/s41588-019-0402-2
| Xyoo | Phau ntawv xov xwm | IF | Lub Npe | Cov ntawv thov |
| 2022 | Kev sib txuas lus ntawm xwm | 17.694 | Lub hauv paus genomic ntawm giga-chromosomes thiab giga-genome ntawm tsob ntoo peony Paeonia ostii | SLAF-GWAS |
| 2015 | Kws Tshawb Fawb Txog Paj Tshiab | 7.433 | Cov cim qhia txog kev yug me nyuam yog cov cim qhia txog cov cheeb tsam genomic uas tseem ceeb rau kev ua liaj ua teb hauv taum pauv | SLAF-GWAS |
| 2022 | Phau Ntawv Xov Xwm Txog Kev Tshawb Fawb Qib Siab | 12.822 | Kev nkag mus rau hauv Gossypium barbadense thoob plaws genome rau hauv G. hirsutum qhia cov loci zoo tshaj plaws rau kev txhim kho tib lub sijhawm ntawm cov paj rwb zoo thiab cov qoob loo cov cwj pwm | SLAF-Kev hloov pauv noob caj noob ces |
| 2019 | Tsob nroj molecular | 10.81 | Kev Tshawb Fawb Txog Cov Neeg thiab Kev Sib Tham De Novo Qhia Txog Keeb Kwm ntawm Weedy Mov ua ib qho kev ua si txog kev hloov pauv | SLAF-Kev hloov pauv noob caj noob ces |
| 2019 | Kev Tshawb Fawb Txog Genes ntawm Xwm | 31.616 | Cov kab ke genome thiab kev sib txawv ntawm cov noob caj noob ces ntawm cov ntses carp, Cyprinus carpio | Daim ntawv qhia SLAF-Linkage |
| 2014 | Kev Tshawb Fawb Txog Genes ntawm Xwm | 25.455 | Cov noob caj noob ces ntawm cov txiv laum huab xeeb cog qoob loo muab kev nkag siab rau hauv legume karyotypes, polyploid kev hloov pauv thiab kev cog qoob loo. | Daim ntawv qhia SLAF-Linkage |
| 2022 | Phau Ntawv Xov Xwm Txog Kev Siv Tshuaj Biotechnology | 9.803 | Kev txheeb xyuas ST1 qhia txog kev xaiv uas cuam tshuam nrog kev caij tsheb ntawm cov noob morphology thiab cov roj thaum lub sijhawm cog taum pauv | Kev tsim cov cim SLAF |
| 2022 | Phau Ntawv Xov Xwm Thoob Ntiaj Teb ntawm Kev Tshawb Fawb Txog Molecular | 6.208 | Kev Txheeb Xyuas thiab Kev Tsim Cov Cim DNA rau Wheat-Leymus mollis 2Ns (2D) Kev Hloov Chaw Chromosome Disomic | Kev tsim cov cim SLAF |
| Xyoo | Phau ntawv xov xwm | IF | Lub Npe | Cov ntawv thov |
| 2023 | Cov ciam teb hauv kev tshawb fawb txog tsob nroj | 6.735 | QTL mapping thiab transcriptome kev tshuaj xyuas ntawm cov suab thaj thaum lub sijhawm txiv hmab txiv ntoo ntawm Pyrus pyrifolia ripening | Daim Ntawv Qhia Txog Genetic |
| 2022 | Phau Ntawv Xov Xwm Txog Kev Siv Tshuaj Biotechnology | 8.154 | Kev txheeb xyuas ST1 qhia txog kev xaiv uas cuam tshuam nrog kev caij tsheb ntawm cov noob morphology thiab cov roj thaum lub sijhawm cog taum pauv
| SNP hu xov tooj |
| 2022 | Cov ciam teb hauv kev tshawb fawb txog tsob nroj | 6.623 | Kev Tshawb Fawb Txog Genome-Wide Association Mapping ntawm Hulless Barely Phenotypes hauv Kev Kub Ntxhov Ib puag ncig.
| GWAS |
