Dear Vangelis, tomorrow Ioannis Karakasiliotis will pass by discussing his run342 As he tried to validate several expressions with qPCR without success I checked his results and found some evidence for potentila sample swaps. The suspicious sampeles are: YK3R3-5291-4aplus YK3R4-5291-4aminus YK3R5-5291-4aminus YK3R9-5269-4fminus YK3R10-5269-4fminus Can you please have a quick check that everything went well with these from your side? Thanks, Martin Dear Ioannis, as already mentioned on the phone, with the modifications for 3UTRseq Panagiotis Moulos made for his metaseqR package, the stranded sequencing data was unfortunately treated as unstranded. The difference in the result is minor, please check below if this improves the verifications you have made. It would help if you could send the names of the genes you selected for verification. Sorry for the inconvenience, but we just discovered this problem recently. Best wishes, Martin At http://genomics-lab.fleming.gr/fleming/external/Karakasiliotis/run342/metaseqr_run_run342a/index.html you will find the metaseqR [1] differential gene expression analysis using the contrasts "Grp4aMinus_vs_Grp4aPlus", "Grp4fMinus_vs_Grp4fPlus" At http://genomics-lab.fleming.gr/fleming/external/Karakasiliotis/run342/metaseqr_run_run342b/index.html you will find the metaseqR differential gene expression analysis using the contrasts "Grp4aMinus4fMinus_vs_Grp4aPlus", "Grp4aMinus4fMinus_vs_Grp4fPlus", "Grp4aMinus4fMinus_vs_Cplus1Plus" At http://genomics-lab.fleming.gr/fleming/external/Karakasiliotis/run342/metaseqr_run_run342d/index.html you will find the metaseqR differential gene expression analysis using the contrast "Cplus_vs_vectorhuh75" from: Martin Reczko to: Ioannis Karakasiliotis cc: Ioannis Karakasiliotis Dear Ioannis, at http://genomics-lab.fleming.gr/cgi-bin/hgTracks?db=mm10&hubUrl=http://genomics-lab.fleming.gr/fleming/external/Karakasiliotis/run342/hub-mmu.txt you will find tracks for the mmu samples in run342, normalized to the smallest library, YK3R8-5269-4fplus with 1659156 reads: sample n_reads YK3R1-5291-4aplus 2738262 YK3R2-5291-4aplus 2271924 YK3R3-5291-4aplus 1941843 YK3R4-5291-4aminus 1888433 YK3R5-5291-4aminus 3164562 YK3R6-5269-4fplus 2943172 YK3R7-5269-4fplus 2097718 YK3R8-5269-4fplus 1659156 YK3R9-5269-4fminus 3179846 YK3R10-5269-4fminus 1995501 YK3R11-Cplus-1plus 2598127 At http://genomics-lab.fleming.gr/cgi-bin/hgTracks?db=hg19&hubUrl=http://genomics-lab.fleming.gr/fleming/external/Karakasiliotis/run342/hub.txt you will find tracks for the hsa samples in run342, normalized to the smallest library, YK3R8-5269-4fplus with 1659156 reads: sample n_reads YK3R12-Cplus-shorthuh75 2282534 YK3R13-vectorhuh75 2334101 At http://genomics-lab.fleming.gr/fleming/external/Karakasiliotis/run342/metaseqr_quantseq_run342a/index.html you will find the metaseqR [1] differential gene expression analysis using the contrasts "Grp4aMinus_vs_Grp4aPlus", "Grp4fMinus_vs_Grp4fPlus" (note the used naming convention: in A_vs_B, A is always the reference). At http://genomics-lab.fleming.gr/fleming/external/Karakasiliotis/run342/metaseqr_quantseq_run342b/index.html you will find the metaseqR differential gene expression analysis using the contrasts "Grp4aMinus4fMinus_vs_Grp4aPlus", "Grp4aMinus4fMinus_vs_Grp4fPlus", "Grp4aMinus4fMinus_vs_Cplus1Plus" At http://genomics-lab.fleming.gr/fleming/external/Karakasiliotis/run342/metaseqr_quantseq_run342c/index.html you will find the metaseqR differential gene expression analysis using the contrast "Cplus_vs_vectorhuh75" Let me know if you need any additional information, best wishes Martin [1] Moulos P and Hatzis P “Systematic integration of RNA-Seq statistical algorithms for accurate detection of differential gene expression patterns.” Nucl. Acids Res. (2014) doi: 10.1093/nar/gku1273 http://nar.oxfordjournals.org/content/early/2014/12/01/nar.gku1273.abstract. #@ 06042017 Hi Martin, For Ion 342 please proceed with the following groups/comparisons: Mouse samples Group 1 (4a) YK3R1_5291 4a + YK3R2_5291 4a + YK3R3_5291 4a + Group 2 (4a Ctrl) YK3R4_5291 4a - YK3R5_5291 4a - Group 3 (4f) YK3R6_5269 4f + YK3R7_5269 4f + YK3R8_5269 4f + Group 4 (4f Ctrl) YK3R9_5269 4f - YK3R10_5269 4f - Group 5 YK3R11_C+1+ ----------------------------------- Group 1 vs 2, Group 3 vs 4, Group 1 vs 2+4, Group 3 vs 2+4, Group 5 vs 2+4. ------------------------------------ Human samples YK3R12_c+1 short huh7.5 vs YK3R13_vector huh7.5 If you need anything else contact me, Vaggelis -- Vaggelis Harokopos, PhD ------------------------------------------------------------------------- Genomics Facility BSRC Al. Fleming 34 Fleming str • Vari • Athens • 16672 • Greece T +30 210 9656310, int. 221 harokopos@fleming.gr • genomics@fleming.gr www.fleming.gr