参数< -列表(缓存= FALSE) # #设置,包括= FALSE ----------------------------------------------------- 美元knitr: opts_chunk设置(图。width=7, fig.height=6) ## ----load, message=FALSE, warning=FALSE--------------------------------------- library(topconfects) library(nbpsq) library(edgeR) library(limma) library(dplyr) library(ggplot2) data(arab) #为每个基因检索符号info <- AnnotationDbi::select(org. at . taair .db::org. at . taair .db, rownames(arab)，"SYMBOL") %>% group_by(TAIR) %>% summarize(SYMBOL=paste(unique(na. ignore (SYMBOL))，collapse="/")) arab_info <- info[match(rownames(arab)，info$TAIR)，] %>% select(-TAIR) %>% as.data.frame rownames(arab_info) <- rownames(arab) #从样本名称中提取实验设计Treat <- factor(substring(colnames(arab)，1,4)) %>% relevel(ref="mock") Time <- factor(substring(colnames(arab)，5,5)) y <- DGEList(arab，基因=as.data.frame(arab_info)) #保留至少3个样本的RPM大于2 Keep <- rowsum (cpm(y)>2) >= 3 y <- y[Keep，， Keep .lib. txt]大小= FALSE) y < - calcNormFactors (y) # #——limma -------------------------------------------------------------------- 设计< - model.matrix(~时间+治疗)设计[,]符合< -轰(y,设计)% > % lmFit(设计)# #——limma_confects ----------------------------------------------------------- 西点< - limma_confects(健康,系数=“Treathrcc”,罗斯福= 0.05)混合# #——无花果。身高= 7 ------------------------------------------------------------- confects_plot(西点 ) ## ----------------------------------------------------------------------------- confects_plot_me(西点 ) ## ---- fig.height = 7 ------------------------------------------------------------ fit_eb < - ebay () < - topTable (fit_eb系数=“Treathrcc”,n =正)rank_rank_plot(糖果表名称,美元rownames(上),“limma_confects”,“topTable ") ## ----------------------------------------------------------------------------- plotMD(健康,传说=“bottomleft”状态= paste0 (ifelse (rownames(适合)% % rownames(上)[1],“topTable”," "),ifelse (rownames(适合)% %糖果表名称[1],美元“西点 ",""))) ## ---- 刨边机 -------------------------------------------------------------------- y < - estimateDisp (y,设计、健壮的= TRUE) efit < glmQLFit (y,设计,健壮的= TRUE) # #——edger_confects ----------------------------------------------------------- econfects < - edger_confects (efit系数=“Treathrcc”,罗斯福= 0.05,= 0.05)步econfects # #——无花果。身高= 7 ------------------------------------------------------------- confects_plot econfects confects_plot_me (econfects ) ## ----------------------------------------------------------------------------- etop修建< - glmQLFTest (efit系数= " Treathrcc ") % > % topTags (n =正)plotMD (efit传奇=“bottomleft”状态= paste0 (ifelse (rownames (efit) % % econfects表名称[1],美元“糖果”," "),ifelse (rownames (efit) % % rownames (etop修建)[1],”topTags ",""))) ## ---- 消息= F,警告= F ----------------------------------------------------- 库(DESeq2) dds < - DESeqDataSetFromMatrix (countData =阿拉伯colData = data.frame(时间、治疗),rowData = arab_info设计= ~时间+治疗)dds < - DESeq (dds ) ## ----------------------------------------------------------------------------- dconfects < - deseq2_confects (name = " Treat_hrcc_vs_mock dds,步骤= 0.05 ) ## ----------------------------------------------------------------------------- < - lfcShrink萎缩(dds, coef="Treat_hrcc_vs_mock", type="ashr") dconfects$table$shrunk <- shrunk$log2FoldChange[dconfects$table$index] dconfects ## ----------------------------------------------------------------------------- table(dconfects$table$filtered) tail(dconfects$table) ## ----fig.height=7------------------------------------------------------------- confects_plot(dconfects) + geom_point(aes(x=shrunk, size=baseMean, color="lfcShrink"), alpha=0.75) ## ----------------------------------------------------------------------------- filter(dconfects$table, !filtered) %>% ggplot(aes( x=ifelse(is.na(confect),0,confect), y=shrunk, color=!is.na(confect))) + geom_point() + geom_abline() + coord_fixed() + theme_bw() + labs(color="Significantly\nnon-zero at\nFDR 0.05", x="confect", y="lfcShrink using ashr") ## ----fig.height=7------------------------------------------------------------- rank_rank_plot(confects$table$name, econfects$table$name, "limma confects", "edgeR confects") rank_rank_plot(confects$table$name, dconfects$table$name, "limma confects", "DESeq2 confects") rank_rank_plot(econfects$table$name, dconfects$table$name, "edgeR confects", "DESeq2 confects") ## ----------------------------------------------------------------------------- sessionInfo()