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Question: Pheatmap annotation

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7.5 years ago

Oliver ▴ 10

Hello,

I am using pheatmap to map RNA-seq data. I have run into the same problem as other users, in attempting to annotate my heatmaps. Unfortunately I'm in the early stages of learning R, so I am struggling to interpret the solutions offered in other posts.

I would like to annotate my heatmap by rows (genes falling into a particular category "Genetype", as well as columns "SubcloneCohort". I get the common error message:

Error in check.length("fill") : 'gpar' element 'fill' must not be length 0

If anyone can help me understand what is amiss I'll be very grateful! Here is a link to the .CSV data file

Thanks, Oliver

The code I am using is as follows:

Read in data

data <- read.csv("DNA repair genes.csv", comment.char="#")

assign labels in column 1 to "rnames"

rnames <- data[,1]

assign labels in column 67 to "gtype"

gtype <- data[,67]

transform column 2-66 into a matrix

matrix <- data.matrix(data[,2:66])
rownames(matrix) <- rnames

Generate annotations for columns

annotation_col = data.frame( SubcloneCohort = factor(rep(c("J", "O", "P", "K", "L", "N", "Q"), c(8, 10, 9, 10, 10, 7, 11))))

rownames(annotation_col) = paste("Cohort", 1:65, sep = "")

Generate annotations for rows

annotation_row = data.frame( GeneType = factor(rep(c("Strand break joining", "Base excision repair", "Poly polymerases", "Direct reversal of damage", "Repair of topoisomerase crosslinks", "Mismatch excision repair", "Homologour recombination", "Fanconi anaemia", "Non-homologous end joining"), c(6, 11, 3, 3, 2, 9, 19, 14, 7))))

rownames(annotation_row) = paste("Gene", 1:74, sep = "")

Display column annotations

pheatmap(matrix, annotation_col = annotation_col)

Draw heatmaps

pheatmap(matrix, cluster_rows=T, cluster_cols=F, scale = "row", clustering_distance_rows = "euclidean", gaps_col = c(8, 18, 27, 37, 47, 54), border_color = FALSE, color = colorRampPalette(c("red", "black", "green"))(499), #add this for special color pallette cellwidth = 8, cellheight = 10, main = "DNA repair", fontsize = 12, fontsize_row = 8, fontsize_col = 7)

pheatmap annotation RNA-Seq • 8.4k views

ADD COMMENT • link 7.5 years ago by Oliver ▴ 10

score 1 · Answer 1 · 2017-05-30

Solved! Updated code below:

#################################

Designate column and row titles

rnames <- data[,1] # assign labels in column 1 to "rnames" cnames <- data[1:65, 68] # assign labels in column 68 to "cnames"

Create matrix from data

matrix <- data.matrix(data[, 2:66]) # transform column 2-67 into a matrix rownames(matrix) <- rnames colnames(matrix) <- cnames

Generate annotations for columns

column_annotations = data.frame( SubcloneCohort = factor(rep(c("MShef4 normal", "MShef11 normal O", "MShef11 normal P", "MShef11 +Y27632 K", "MShef11 +Y27632 L", "MShef11 5% oxygen N", "MShef11 5% oxygen Q"), c(8, 10, 9, 10, 10, 7, 11))))

rownames(column_annotations) = colnames(matrix)

Generate annotations for rows

row_annotations = data.frame( GeneType = factor(rep(c("Strand break joining", "Base excision repair", "Poly polymerases", "Direct reversal of damage", "Repair of topoisomerase crosslinks", "Mismatch excision repair", "Homologour recombination", "Fanconi anaemia", "Non-homologous end joining"), c(6, 11, 3, 3, 2, 9, 19, 14, 7))))

rownames(row_annotations) = rownames(matrix)

Pheatmap drawing of heatmap

Draw heatmaps with annotation, fix cell sizes, and save to file with correct size

pheatmap(matrix, cluster_rows=T, cluster_cols=F, scale = "row", clustering_distance_rows = "correlation", annotation_row = row_annotations, annotation_col = column_annotations, annotation_legend = TRUE, gaps_col = c(8, 18, 27, 37, 47, 54), border_color = FALSE, color = colorRampPalette(c("red", "black", "green"))(499), #add this for special color pallette cellwidth = 6, cellheight = 6, main = "DNA repair genes", fontsize = 8, fontsize_row = 6, fontsize_col = 7)

dev.off()