Deseq2: 3 factors design

Question

[marburg2107]

It's really complicated for me to handle three-factors with DESeq2, though many of the posts on Biostars.I think the developer should make it more clear to users.

Three factors are temperature(A, B); water content (C, D); co2 concentration (E, F).

I want to know the following effects: (1)temperature, (2)water, (3)co2, (4)temperatureco2, (5)waterco2, (6)temperaturewaterco2, (7)temperature*water.

I have tried a lot with design in Deseq2, like DESeqDataSetFromMatrix(countData = countData,colData = colData,design = ~co2+temperature+water+co2:temperature+co2:water)...but no design can tell me temperaturewaterco2 effect.

Thanks!

Answer 1

[marburg2107]

countData = read.table ("C:/Users/bei/Desktop/deseq2/kegg.txt", sep= "\t", row.names = 1, head = TRUE)
colData = read.table ("C:/Users/bei/Desktop/deseq2/151617-new2.txt", sep= "\t", row.names = 1, header = TRUE)
colData[['co2']] = factor(colData[['co2']], levels = c('TRUE', 'FALSE'))
colData[['warm']] = factor(colData[['warm']], levels = c('TRUE', 'FALSE'))>
colData[['drought']] = factor(colData[['drought']], levels = c('TRUE', 'FALSE'))
dataset <- DESeqDataSetFromMatrix(countData = countData,colData = colData,design = ~ co2 + warm+ drought)
dds <- DESeq(dataset)
resultsNames(dds)
results(dds, contrast=list(c("co2_FALSE_vs_TRUE", "warm_FALSE_vs_TRUE", "drought_FALSE_vs_TRUE")))

Is it right to calculated the effect of co2droughtwarm?

Design matrix is:

sample co2 warm drougt
1 TRUE TRUE TRUE
2 TRUE FALSE FALSE
3 FALSE TRUE FALSE

...

Answer 2

[marburg2107]

DESeqDataSetFromMatrix(countData = countData,colData = colData,design = ~co2+warm+drought+co2:warm+co2:drought+warm:drought+co2:warm:drought) --------------Error in checkFullRank(modelMatrix)

My R version is 3.4.2 (2017-09-28), Deseq2 version is 1.18.1

[russhh]

could you post the contents of your design matrix

[marburg2107]

Total 72 samples, 6 treatment,12 replicates.

6 treatments were: co2-400ppm, co2-400ppm+drought, co2-400ppm+drought+warm, co2-600ppm, co2-600ppm+drought,co2-600ppm+drought+warm

[russhh]

the actual design matrix. From the description in the first post you've got 2^3 = 8 treatments. Please. The actual design matrix

[marburg2107]

The design of the experiment looks like:

sample co2 warm drought
a11 400ppm yes 125mm
a12 400ppm yes 125mm
a13 400ppm yes 125mm
a14 400ppm yes 125mm
a21 400ppm yes 125mm
a22 400ppm yes 125mm
a23 400ppm yes 125mm
a24 400ppm yes 125mm
a31 400ppm yes 125mm
a32 400ppm yes 125mm
a33 400ppm yes 125mm
a34 400ppm yes 125mm
e11 480ppm yes 125mm
e12 480ppm yes 125mm
e13 480ppm yes 125mm
e14 480ppm yes 125mm
e21 480ppm yes 125mm
e22 480ppm yes 125mm
e23 480ppm yes 125mm
e24 480ppm yes 125mm
e31 480ppm yes 125mm
e32 480ppm yes 125mm
e33 480ppm yes 125mm
e34 480ppm yes 125mm
A11 400ppm no 125mm
A12 400ppm no 125mm
A13 400ppm no 125mm
A14 400ppm no 125mm
A21 400ppm no 125mm
A22 400ppm no 125mm
A23 400ppm no 125mm
A24 400ppm no 125mm
A31 400ppm no 125mm
A32 400ppm no 125mm
A33 400ppm no 125mm
A34 400ppm no 125mm
E11 480ppm no 125mm
E12 480ppm no 125mm
E13 480ppm no 125mm
E14 480ppm no 125mm
E21 480ppm no 125mm
E22 480ppm no 125mm
E23 480ppm no 125mm
E24 480ppm no 125mm
E31 480ppm no 125mm
E32 480ppm no 125mm
E33 480ppm no 125mm
E34 480ppm no 125mm
aa11 400ppm no 250mm
aa12 400ppm no 250mm
aa13 400ppm no 250mm
aa14 400ppm no 250mm
aa21 400ppm no 250mm
aa22 400ppm no 250mm
aa23 400ppm no 250mm
aa24 400ppm no 250mm
aa31 400ppm no 250mm
aa32 400ppm no 250mm
aa33 400ppm no 250mm
aa34 400ppm no 250mm
ee11 480ppm no 250mm
ee12 480ppm no 250mm
ee13 480ppm no 250mm
ee14 480ppm no 250mm
ee21 480ppm no 250mm
ee22 480ppm no 250mm
ee23 480ppm no 250mm
ee24 480ppm no 250mm
ee31 480ppm no 250mm
ee32 480ppm no 250mm
ee33 480ppm no 250mm
ee34 480ppm no 250mm

[marburg2107]

1. DESeqDataSetFromMatrix(countData = countData,colData = colData,design = ~ co2 + warm+ drought+co2:warm+co2:drought) (no problem)

2. DESeqDataSetFromMatrix(countData = countData,colData = colData,design = ~ co2 + warm+ drought+co2:warm+co2:drought+co2:warm:drought)(the model matrix is not full rank)

[marburg2107]

Thanks for your help!

I just have 6 treatments as co2(400ppm),co2(480ppm), co2(400ppm)+drought(125mm), co2(480ppm)+drought(125mm),co2(400ppm)+drought(125mm)+warm(yes),co2(400ppm)+drought(125mm)+warm(no).

As you said, we should have 8 treatments. BUT they did not do other treatments.

I want to know the following effects: (1)warm, (2)drought, (3)co2, (4)warm+co2, (5)drought+co2, (6)warm+drought+co2, (7)warm+drought from above 6 treatments.