Microarray DEG scatterplot

Question

[bioinformatics]

Hi,

I have found that my selected gene, probe I.D 201667_at is differentially expressed between WDLPS and DDLPS tumour tissue samples after performing microarray DEG analysis.

Instead of just a p value in a table format:

Probe I.D "201667_at" logFC 10.8205874181535 AveExpr 10.6925705768407 t 82.8808890739766 P.Value 3.10189446528995e-88 adj.P Val 3.10189446528995e-88 "B" 191.589248589131

I have decided to present the data as a scatter plot/plot MDS (with an error bar) using expression values of the specific gene between the two tumour types (40 vs 52 samples) to show that it is differentially expressed. So 92 dots/points in total.

The commands I used for microarray differential expression analysis are:

library("arrayQualityMetrics")
library(GEOquery)
library(oligo)
library(Biobase)
library(affy)
library("splitstackshape")
library("tidyr")
library("dplyr")

celFiles <- list.celfiles()
affyRaw <- oligo::rma(affyraw)
eset <- oligo::rma(affyRaw)
library(limma)
pData(eset)
Groups <- c("DDLPS", "DDLPS", "WDLPS", "WDLPS")
design <- model.matrix(~factor(Groups))
colnames(design) <- c("DDLPS", "DDLPSvsWDLPS")
fit <- lmFit(eset, design)
fit <- eBayes(fit)
option (digits =2)
res <- topTable (fit, number = Inf, adjust.method = "none", coef = 1)
write.table(res, "diff_exp.txt", sep= "\t)
require(hgu133a.db)
annotLookup <- select(hgu133a.db, keys = probes,
  columns = c('PROBEID', 'ENSEMBL', 'SYMBOL'))

content <- read_csv(path, col_names = TRUE)

path <- "/Users/mesalie/expressionvalues.csv"
content <- read_csv(path, col_names = TRUE)

ggplot(content, aes(x = log(DDLPS, WDLPS)), y = log(Epression value2, Expression value4))) + geom_point(aes(color= factor(gear)))

Error in factor(gear) : object 'gear' not found However I keep getting the following error message for the ggplot function.

the list content is below head (content)

# A tibble: 6 × 4
  DDLPS         `Expression value...2` WDLPS         `Expression value...4`
  <chr>                          <dbl> <chr>                          <dbl>
1 GSM766533.CEL                  10.0  GSM766621.CEL                  11.6 
2 GSM766534.CEL                   9.29 GSM766622.CEL                  10.1 
3 GSM766535.CEL                  10.8  GSM766623.CEL                   9.87
4 GSM766536.CEL                  10.5  GSM766624.CEL                  10.4 
5 GSM766537.CEL                  10.7  GSM766625.CEL                  10.5 
6 GSM766538.CEL                  10.1  GSM766626.CEL                  10.2 

Can anyone please advise me on how I can create the 'gear' object?

Thankyou

[Basti]

There is no gear variable in your content dataset. Replace it with the name of the variable you want as color on your plot.

[bioinformatics]

I have done this and received the following error message:

ggplot(content, aes(x = log(DDLPS, WDLPS)), y = log(Epression value2, Expression value4))) + geom_point(aes(color= factor(WDLPS, DDLPS))))

Error: unexpected symbol in "ggplot(content, aes(x = log(DDLPS, WDLPS)), y = log(Epression value2"

Do you also know what the unexpected symbol in this command line is?

[Basti]

There are too many brackets in your code line. Please take the time to check the code you run and impregnate about R coding.

[bioinformatics]

Ok thanks for your help! Do you know which brackets to remove?

ggplot(content, aes(x = log(DDLPS, WDLPS)), y = log(Epression value2, Expression value4) + geom_point(aes(color= factor(WDLPS, DDLPS))))

[Basti]

ggplot(content, aes(x = log(DDLPS, WDLPS)), y = log(`Expression value...2`, `Expression value...4`)) +
         geom_point(aes(color= factor(WDLPS, DDLPS)))

Please be careful with every single code you write, there were plenty of errors in your code, variables that were not called by their correct name, brackets placed at wrong position

[bioinformatics]

Thankyou for your corrections.

I used this command:

ggplot(content, aes(x = log(DDLPS, WDLPS)), y = log(Expression value...2, Expression value...4)) + geom_point(aes(color= factor(WDLPS, DDLPS)))

and got the following error message.

Error in log(DDLPS, WDLPS) : non-numeric argument to mathematical function

Do you know what this means?

[Basti]

Do you think it is possible to take the log of a character variable? Because this is what you are trying to do here. I think you are completely lost in anything you are doing here, I suggest to get in touch with someone that can learn you R programming in your lab/school before attempting to do microarray analysis.