|
|
| Line 1: |
Line 1: |
| − | == Ongoing ==
| + | [[2017 Jan Taeyoung Lab note]] |
| − | | + | |
| − | 1. TBLASTX using Jat Species Transcriptome
| + | |
| − | | + | |
| − | == 2017 1.2 ==
| + | |
| − | ==== Jatropha transcriptome Trinity assemble ====
| + | |
| − | raw data : 244:/NGS/NGS/JatrophaCurcas/RNA
| + | |
| − | Jatropha species transcriptome assemble : Jct,Jcu, Jin, Jgo, Jci, Jpo, Jmu, Jma, Jac, Rco (listed in 244:/NGS/NGS/JatrophaCurcas/RNA/list). All done
| + | |
| − | Jatropha organ transcriptome assemble : Leaf, Root, Stem, Female flower, Male flower, LG, SG, Y, B. All done
| + | |
| − | | + | |
| − | ==== Cdhit ====
| + | |
| − | 193:/data2/alima90/program/cdhit/cd-hit -i Y.cds.fa -M 10000 -o Y.cds.fa.cdhit -T 5
| + | |
| − | 193:/data2/alima90/program/cdhit/cd-hit -i LG.cds.fa -M 10000 -o LG.cds.fa.cdhit -T 5
| + | |
| − | | + | |
| − | ==== UV GBS mapping (w/ joinmap) ====
| + | |
| − | 244:python vcf.parsing.for.mandf.py UV.vcf.SNPonly 3 0.01 except_sample.txt > UV.vcf.SNPonly.except.LowDepthSample.d3.Q30.m0.1.loc
| + | |
| − | loc file is manually edited by excel
| + | |
| − | Genetic map is constructed using Joinmap 4.1
| + | |
| − | | + | |
| − | ==== KaKs calculation using scripts provided by MCscanX ====
| + | |
| − | | + | |
| − | '''KaKs calculation between Jatropha species'''
| + | |
| − | 244 :python /alima9002/63_backup/Jatropha/CDS/run.kaks.py
| + | |
| − | | + | |
| − | ==== Large Insertion Prediction ====
| + | |
| − | ===== LIP short primer preparation =====
| + | |
| − | '''Primer info'''
| + | |
| − | >LIP01short_F
| + | |
| − | AACTGAACACAGACAATGAA
| + | |
| − | >LIP01short_R
| + | |
| − | CAATTTATACACCACCTTAC
| + | |
| − | >LIP02short_F
| + | |
| − | CTCTTTGTATTTGGTGACAA
| + | |
| − | >LIP02short_R
| + | |
| − | GTATTAGCAGCTTTTGCTTA
| + | |
| − | >LIP03short_F
| + | |
| − | AATTGTAAGACATATCCCTC
| + | |
| − | >LIP03short_R
| + | |
| − | CTGCCCCACTAATAATTAAT
| + | |
| − | >LIP04short_F
| + | |
| − | TAAAAACAGAACTTGTCCAC
| + | |
| − | >LIP04short_R
| + | |
| − | ATCACAAGACTGAACAAGTA
| + | |
| − | >LIP05short_F
| + | |
| − | ATTGACATAAGGTTGCATAG
| + | |
| − | >LIP05short_R
| + | |
| − | CCTTAGCTCTTTTCTTTTGT
| + | |
| − | >LIP06short_F
| + | |
| − | GAAGGAAGGAAGCAATTATT
| + | |
| − | >LIP06short_R
| + | |
| − | TGACTTACCCTTTTTACCTT
| + | |
| − | >LIP07short_F
| + | |
| − | CACATGTTTGTCACTCTAAT
| + | |
| − | >LIP07short_R
| + | |
| − | GAAGTGAGGCCTAAAATAAA
| + | |
| − | >LIP08short_F
| + | |
| − | GAATGTATTGTCTTTGATCC
| + | |
| − | >LIP08short_R
| + | |
| − | GTTGGATTTTGTTCTTTCCA
| + | |
| − | >LIP09short_F
| + | |
| − | AGAAAAACGTCGATACCAAA
| + | |
| − | >LIP09short_R
| + | |
| − | CGATTTAGTAACCTTAGAAC
| + | |
| − | >LIP10short_F
| + | |
| − | ATCTTCAAAATGTCTCTAGG
| + | |
| − | >LIP10short_R
| + | |
| − | TACAGATATTCTTAGGCAGT
| + | |
| − | >LIP11short_F
| + | |
| − | TGTAACTCTCAATTAAGCAG
| + | |
| − | >LIP11short_R
| + | |
| − | ATCTTTCTGTAAGCACTTAG
| + | |
| − | >LIP12short_F
| + | |
| − | CTAGAACCGATTTGTTCAAA
| + | |
| − | >LIP12short_R
| + | |
| − | GCAGTTGTTTTGGATTAACA
| + | |
| − | >LIP13short_F
| + | |
| − | AAAGAGAAAGCAGAGAAATC
| + | |
| − | >LIP13short_R
| + | |
| − | ATGTATAGATTGGAGGAAAG
| + | |
| − | >LIP14short_F
| + | |
| − | ATTATGGAAAGGAATTGGAG
| + | |
| − | >LIP14short_R
| + | |
| − | CCATGTCTAGTATTTACTCA
| + | |
| − | >LIP15short_F
| + | |
| − | TTAATGACTGATCGTTAGTG
| + | |
| − | >LIP15short_R
| + | |
| − | CGGGAGTTATGAAAAATAGT
| + | |
| − | >LIP24short_F
| + | |
| − | AGTATGGTTTCAACATATGG
| + | |
| − | >LIP24short_R
| + | |
| − | GATATGAAGTTGACATGCTA
| + | |
| − | >LIP16short_F
| + | |
| − | ATTTAAAAGCTCGTAACTCC
| + | |
| − | >LIP16short_R
| + | |
| − | GGATAAGCAATTACAACACA
| + | |
| − | >LIP17short_F
| + | |
| − | CCCAAATTTTTAAATGCACC
| + | |
| − | >LIP17short_R
| + | |
| − | CTCTTGGAACGTGAAAAATT
| + | |
| − | >LIP18short_F
| + | |
| − | TTTTCTAGAAGGATTTGTGC
| + | |
| − | >LIP18short_R
| + | |
| − | CCATGCAAACCCAATTTTAA
| + | |
| − | >LIP19short_F
| + | |
| − | GTAAAACTAAGGTTGAGCTA
| + | |
| − | >LIP19short_R
| + | |
| − | CCACAAGTCACAACAATTTA
| + | |
| − | >LIP20short_F
| + | |
| − | TTATTTGTATGTTGGAGACC
| + | |
| − | >LIP20short_R
| + | |
| − | CATGGTATATAGGTTTAGGT
| + | |
| − | >LIP21short_F
| + | |
| − | CATAGAGAGTTTTGGATTAC
| + | |
| − | >LIP21short_R
| + | |
| − | AAAGAACTGATAGTGTCATG
| + | |
| − | >LIP22short_F
| + | |
| − | ATATGTACATGTATGGTGTG
| + | |
| − | >LIP22short_R
| + | |
| − | CCTAAATCTAGCAGAAGATT
| + | |
| − | >LIP23short_F
| + | |
| − | ATGTATGGAGAAATGGGTTA
| + | |
| − | >LIP23short_R
| + | |
| − | ATATAGAAATGGAGGTTGCT
| + | |
| − | (listed in BACKUP(J:)/박사/Indel Candidate/LIP_short_primer.fa)
| + | |
| − | | + | |
| − | Primer dilution
| + | |
| − | | + | |
| − | == 2017. 1. 3 ==
| + | |
| − | ===AMORE work===
| + | |
| − | | + | |
| − | python ~/py/ret_fasta_by_gene_name.py /alima9002/ref/Athaliana/annotation/Athaliana_167_TAIR10.cds.fa gene_list.txt > gene_list.txt.fa
| + | |
| − | blastp -db /alima9002/ref/Gmax/annotation/Gmax_275_Wm82.a2.v1.protein.fa -query gene_list.txt.pep.fa -evalue 1e-5 -num_alignments 1 -outfmt 6 -num_threads 6 -out gene_list.txt.pep.fablastp.Gm275.1e-5.out6
| + | |
| − | | + | |
| − | '''Homolog with Ath'''
| + | |
| − | Glyma.08G014900
| + | |
| − | Glyma.05G208300
| + | |
| − | Glyma.20G001900
| + | |
| − | Glyma.03G176600
| + | |
| − | Glyma.19G177400
| + | |
| − | Glyma.03G262600
| + | |
| − | Glyma.06G202300
| + | |
| − | Glyma.05G021800
| + | |
| − | Glyma.17G077700
| + | |
| − | Glyma.05G022000
| + | |
| − | Glyma.09G234900
| + | |
| − | Glyma.19G025000
| + | |
| − | Glyma.10G224000
| + | |
| − | Glyma.02G081000
| + | |
| − | Glyma.20G167800
| + | |
| − | Glyma.14G072700
| + | |
| − | Glyma.17G252200
| + | |
| − | Glyma.17G050500
| + | |
| − | Glyma.07G038000
| + | |
| − | Glyma.13G109100
| + | |
| − | Glyma.16G007200
| + | |
| − | Glyma.19G105100
| + | |
| − | Glyma.09G283800
| + | |
| − | Glyma.20G172700
| + | |
| − | Glyma.02G076300
| + | |
| − | | + | |
| − | '''SNP typing among IT182932,IT1099098,Hwangkeum-Kong'''
| + | |
| − | | + | |
| − | 1.Read mapping using bwa mem with default options (/home/hayasen/Workspace/Glycine/GlycineMax/ver275/Reads/)
| + | |
| − | | + | |
| − | 2.mpileup
| + | |
| − | samtools mpileup -f /alima9002/ref/Gmax/assembly/Gmax_275_v2.0.fa -v -t DP,AD,ADF,ADR,SP,INFO/AD,INFO/ADF,INFO/ADR -u -b bam_list | bcftools call -v -m -O v > Variant.vcf
| + | |
| − | | + | |
| − | ===LIP short primer gradient PCR===
| + | |
| − | 1~8 primer is tested with CS-12
| + | |
| − | | + | |
| − | Gradiant lower temp is 50 upper temp is 65
| + | |
| − | | + | |
| − | Sample is loaded on 1% agarose gel and It was run with 100 V on 1 hour.
| + | |
| − | | + | |
| − | {| class="wikitable"
| + | |
| − | |-
| + | |
| − | | 52.7 || 54.1 || 55.5 || 56.8 || 58.2 | 59.5 || 60.9 || 62.3
| + | |
| − | |}
| + | |
| − | | + | |
| − | <gallery>
| + | |
| − | File:17010301.jpeg|LIPshort01-04
| + | |
| − | </gallery>
| + | |
| − | 1 2
| + | |
| − | | + | |
| − | 3 4
| + | |
| − | | + | |
| − | | + | |
| − | LIPshort1 -> Error when it is loaded
| + | |
| − | | + | |
| − | LIPshort4 -> 55.5~59.5에서 증폭한 샘플만 로딩
| + | |
| − | | + | |
| − | <gallery>
| + | |
| − | File:17010302.jpg|Caption2
| + | |
| − | </gallery>
| + | |
| − | | + | |
| − | 5 6
| + | |
| − | | + | |
| − | 7 8
| + | |
| − | | + | |
| − | Estimated Tm:55.5~56.8
| + | |
| − | | + | |
| − | | + | |
| − | == 2017.1.4~2017.1.6 ==
| + | |
| − | | + | |
| − | 농장 출장
| + | |
| − | | + | |
| − | ===LIP Gradient PCR===
| + | |
| − | 50~65 celsius degree
| + | |
| − | | + | |
| − | 1% agar 100V 1h
| + | |
| − | | + | |
| − | | + | |
| − | | + | |
| − | <gallery>
| + | |
| − | File:2017010401.jpg|LIP01,09,10,11
| + | |
| − | File:2017010402.jpg|LIP12,13,14,15
| + | |
| − | </gallery>
| + | |
| − | | + | |
| − | All good
| + | |
| − | | + | |
| − | | + | |
| − | <gallery>
| + | |
| − | File:2017010601.jpg|LIP16,17,18,19
| + | |
| − | File:2017010602.jpg|LIP20,21,22,24
| + | |
| − | </gallery>
| + | |
| − | | + | |
| − | LIP16's lower band is our target
| + | |
| − | | + | |
| − | LIP20 did not show band
| + | |
| − | | + | |
| − | == 2017.1.9 ==
| + | |
| − | | + | |
| − | ===AMORE(GK, IT182932, IT109098)===
| + | |
| − | ==== VCF parsing ====
| + | |
| − | python ~/py/Reseq/filter.vcf.by.phred.hetero.depth.py Variant.vcf.SNP 5 > Variant.vcf.SNP.filtered.d5.Q30.homo
| + | |
| − | ==== Typing ====
| + | |
| − | cat Variant.vcf.SNP.filtered.d5.Q30.homo.diff | python ~/py/Reseq/[Reseq]SNP_counter.py /alima9002/ref/Gmax/annotation/Gmax_275_Wm82.a2.v1.gene_exons.gff3 /alima9002/ref/Gmax/assembly/Gmax_275_v2.0.fa 30 > Variant.vcf.SNP.filtered.d5.Q30.homo.diff.type
| + | |
| − | | + | |
| − | ===Jatropha OrthoMcl===
| + | |
| − | Retrieve complete pep only for OrthoMcl
| + | |
| − | | + | |
| − | == 2017.1.10 ==
| + | |
| − | ===AMORE(GK, IT182932, IT109098)===
| + | |
| − | ==== filtering SNPs on homologous ====
| + | |
| − | python ret_homolog_only.py Variant.vcf.SNP.filtered.d5.Q30.homo.diff.type homologs.txt > Variant.vcf.SNP.filtered.d5.Q30.homo.diff.type.homologs.only
| + | |
| − | ==== Syn or Nonsyn typing ====
| + | |
| − | python ~/py/Reseq/\[Reseq\]det_syn.py /alima9002/ref/Gmax/annotation/Gmax_275_Wm82.a2.v1.gene_exons.gff3 /alima9002/ref/Gmax/assembly/Gmax_275_v2.0.fa Variant.vcf.SNP.filtered.d5.Q30.homo.diff.type.homologs.only.CDS > Variant.vcf.SNP.filtered.d5.Q30.homo.diff.type.homologs.only.CDS.SynNonsyn
| + | |
| − | ===Lactuca Indica Cdhit===
| + | |
| − | /alima9002/program/cd-hit-v4.6.4-2015-0603/cd-hit -i L.Trinity.fasta -o L.Trinity.fasta.cdhit -T 10 -M 10000
| + | |
| − | | + | |
| − | == 2017.1.11==
| + | |
| − | 농장 출장
| + | |
| − | | + | |
| − | == 2017.1.12==
| + | |
| − | ===Amore===
| + | |
| − | ====Make SNP tables====
| + | |
| − | ====INDEL analysis using snpEff====
| + | |
| − | java -jar /alima9002/program/snpEff/snpEff.jar ann -c /alima9002/program/snpEff/snpEff.config -ud 1000 gmax275 Variant.vcf.INDEL > Variant.vcf.INDEL.snpEff
| + | |
| − | | + | |
| − | ==== homologs filtering using annotation file ====
| + | |
| − | ==== VCF filter by homologs retrieved by ann file ====
| + | |
| − | python ret_homolog_only.py Variant.vcf.SNP.filtered.d5.Q30.homo.diff.type homologs.by.ann.txt > Variant.vcf.SNP.filtered.d5.Q30.homo.diff.type.hom.ann
| + | |
| − | ==== Determination Synonymous ====
| + | |
| − | python ~/py/Reseq/\[Reseq\]det_syn.py /alima9002/ref/Gmax/annotation/Gmax_275_Wm82.a2.v1.gene_exons.gff3 /alima9002/ref/Gmax/assembly/Gmax_275_v2.0.fa Variant.vcf.SNP.filtered.d5.Q30.homo.diff.type.hom.ann.CDS > Variant.vcf.SNP.filtered.d5.Q30.homo.diff.type.hom.ann.CDS.SynNonsyn
| + | |
| − | | + | |
| − | ===Jatropha KaKs===
| + | |
| − | python parsing.all.kaks.py all.kaks > all.kaks.ksonly
| + | |
| − | ====Drawing graph using R====
| + | |
| − | require(ggplot2)
| + | |
| − | data<-read.table("all.kaks.ksonly",header=F)
| + | |
| − | colnames(data)<-c("Species","Ks")
| + | |
| − | Ks <- data$Ks
| + | |
| − | Species <- data$Species
| + | |
| − | ggplot(data,aes(Ks,colour=Species))+geom_freqpoly(binwidth=0.01)+scale_x_continuous(limits=c(0,0.8))
| + | |
| − | | + | |
| − | ==2017.1.13==
| + | |
| − | 농장 출장(꼬투리 lwt)
| + | |
| − | | + | |
| − | ==2017.1.16==
| + | |
| − | ===Jatropha Ks value using transcriptome===
| + | |
| − | Jat species which were not clustered by cdhit were used for TBLASTX
| + | |
| − | tblastx -db Jct.cds.fa.complete.fa -query Jgo.cds.fa.complete.fa -evalue 1e-10 -outfmt 6 -num_alignments 5 -out Jct.tblastx.nocdhit.Jgo.1e-10.out6 -num_threads 8
| + | |
| − | | + | |
| − | ===Amore snpEff===
| + | |
| − | Split result as one by one lines
| + | |
| − | perl /alima9002/program/snpEff/scripts/vcfEffOnePerLine.pl Variant.vcf.INDEL.snpEff
| + | |
| − | | + | |
| − | ===Amore SNP typing===
| + | |
| − | for check IT182932 mapping depth, SNP typing is performed in not filtered vcf file
| + | |
| − | python ~/py/Reseq/\[Reseq\]SNP_counter.py Variant.vcf.SNP /alima9002/ref/Gmax/assembly/Gmax_275_v2.0.fa 30 > Variant.vcf.SNP.type
| + | |
| − | | + | |
| − | ==2017.1.17==
| + | |
| − | ===OrthoMcl for Jat Organ===
| + | |
| − | blastp -db goodProteins.fasta -query goodProteins.fasta -outfmt 6 -out goodProteins.fasta.allvall.jat.organ -num_threads 15 -evalue 1e-5 -seg yes -soft_masking true -max_target_seqs 999999999
| + | |
| − | ===Drawing Venn diagram using JatSp orthomcl result===
| + | |
| − | D:\Lab work\Jatropha\JatSp_Orthomcl_Venn
| + | |
| − | ===Re-SNP typing of amore study===
| + | |
| − | 명령어가 잘못된 것을 발견
| + | |
| − | cat Variant.vcf.SNP | python ~/py/Reseq/\[Reseq\]SNP_counter.py /alima9002/ref/Gmax/annotation/Gmax_275_Wm82.a2.v1.gene_exons.gff3 /alima9002/ref/Gmax/assembly/Gmax_275_v2.0.fa 30 > Variant.vcf.SNP.type
| + | |
| − | ===snpEff parsing===
| + | |
| − | 명령어가 잘못된 것을 발견
| + | |
| − | cat Variant.vcf.INDEL.snpEff | 'perl' /alima9002/program/snpEff/scripts/vcfEffOnePerLine.pl > Variant.vcf.INDEL.snpEff.parsed
| + | |
| − | | + | |
| − | ===snpEff typing===
| + | |
| − | python Indel.Typing.Using.snpEff.Result.py Variant.vcf.INDEL.snpEff.parsed > Variant.vcf.INDEL.snpEff.parsed.tp
| + | |
| − | | + | |
| − | ===filtering homologs only on snpEff typing results===
| + | |
| − | python get_INDEL_on_homolog.py Variant.vcf.INDEL.snpEff.parsed.tp homologs.by.ann.txt > Variant.vcf.INDEL.snpEff.parsed.tp.hom.ann
| + | |
| − | less Variant.vcf.INDEL.snpEff.parsed.tp.hom.ann | sort -u > Variant.vcf.INDEL.snpEff.parsed.tp.hom.ann.sorted
| + | |
| − | | + | |
| − | ===count INDEL type===
| + | |
| − | python count.indel.component.py Variant.vcf.INDEL.snpEff.parsed.tp.hom.ann.sorted
| + | |
| − | | + | |
| − | ===Discussion===
| + | |
| − | organ은 3반복을 따로 orthomcl에 집어넣고 셋 다 있는 것을 카운트
| + | |
| − | | + | |
| − | ==2017.1.18==
| + | |
| − | ===ret SNP sets(1 missing GT is permitted)===
| + | |
| − | python filter.vcf.by.phred.hetero.depth.py.only.one.missing.permitted.py
| + | |
| − | ===Amore SNP typing(1 missing GT is permitted)===
| + | |
| − | cat Variant.vcf.SNP.filtered.d5.Q30.missing1.diff | python ~/py/Reseq/\[Reseq\]SNP_counter.py /alima9002/ref/Gmax/annotation/Gmax_275_Wm82.a2.v1.gene_exons.gff3 /alima9002/ref/Gmax/assembly/Gmax_275_v2.0.fa 30 > Variant.vcf.SNP.filtered.d5.Q30.missing1.diff.type
| + | |
| − | | + | |
| − | ==2017.1.19==
| + | |
| − | ===LIP related figure===
| + | |
| − | LIP로 예측한 insertion type별 그림
| + | |
| − | python ~/py/GeneInfo2Figure_v1.1.py gene_list.gi Glyma.08G.123500.gi2f.config > gene_list.gi.svg
| + | |
| − | 이후 Illustrator로 manually 수정
| + | |
| − | | + | |
| − | ==2017.1.23==
| + | |
| − | ===finishing SNP, INDEL typing of amore study===
| + | |
| − | ===Jat species with no cdhit OrthoMcl ===
| + | |
| − | follow Orthomcl manual
| + | |
| − | | + | |
| − | ==2017.1.31==
| + | |
| − | ===Drawing size distribution of predicted insertion ===
| + | |
| − | R with ggplot2 ver. 2.2
| + | |
| − | ggplot(SV, aes(SV$Size))+geom_histogram(aes(fill=SV$Program),binwidth=200,position="dodge")+scale_y_continuous("Counts")+coord_cartesian(ylim=c(400,2000),expand=FALSE)
| + | |
