normilization with RMA, human versus citrus

Question

[zizigolu]

Hi friends, I need your help please. I tried to adopt a tutorial in human (GSE27447) to my case in citrus sinensis, I searched a lot in google but I could not find fit equal to some parts and when I did these steps by removing parts I had doubts about, the result was a rma.txt which could not be read in some case (with ????) I do not know what I should type instead of... my query is GSE67376 (citrus sinensis)

source("http://bioconductor.org/biocLite.R")

biocLite("GEOquery")
biocLite("affy")
biocLite("gcrma")
biocLite("hugene10stv1cdf") # changed to biocLite("citruscdf")
biocLite("hugene10stv1probe") # changed to biocLite("citrusprobe")
biocLite("hugene10stprobeset.db") ????????????
biocLite("hugene10sttranscriptcluster.db") ???????

library(GEOquery)
library(affy)
library(gcrma)
library(hugene10stv1cdf) # changed to library("citruscdf")
library(hugene10stv1probe) # changed to library("citrusprobe")
library(hugene10stprobeset.db) ?????????
library(hugene10sttranscriptcluster.db) ?????

setwd("C:/Users/Man/Desktop/New folder (3)")

getGEOSuppFiles("GSE27447")

setwd("C:/Users/Man/Desktop/New folder (3)/GSE27447")

untar("GSE27447_RAW.tar", exdir="data")

cels = list.files("data/", pattern = "CEL")
sapply(paste("data", cels, sep="/"), gunzip)
cels = list.files("data/", pattern = "CEL")

setwd("C:/Users/Man/Desktop/New folder (3)/data")
raw.data=ReadAffy(verbose=TRUE, filenames=cels, cdfname="hugene10stv1") ??????

data.rma.norm=rma(raw.data)
rma=exprs(data.rma.norm)
rma=format(rma, digits=5)

ls("package:hugene10stprobeset.db") #Annotations at the exon probeset level ?????????
ls("package:hugene10sttranscriptcluster.db") ??????????????

probes=row.names(rma)
Symbols = unlist(mget(probes, hugene10sttranscriptclusterSYMBOL, ifnotfound=NA)) ???????????
Entrez_IDs = unlist(mget(probes, hugene10sttranscriptclusterENTREZID, ifnotfound=NA)) ????????

rma=cbind(probes,Symbols,Entrez_IDs,rma)
write.table(rma, file = "rma.txt", quote = FALSE, sep = "\t", row.names = FALSE, col.names = TRUE)

Answer 1

[Deepak Tanwar]

biocLite("hugene10stprobeset.db") ????????????
biocLite("hugene10sttranscriptcluster.db") ???????

library(hugene10stprobeset.db) ?????????
library(hugene10sttranscriptcluster.db) ?????

These lines do nothing. I don't think that there is a package called citrus.db.

raw.data=ReadAffy(verbose=TRUE, filenames=cels, cdfname="hugene10stv1") ??????
raw.data=ReadAffy(verbose=TRUE, filenames=cels, cdfname="citruscdf")

ls("package:hugene10stprobeset.db") #Annotations at the exon probeset level?????????
ls("package:hugene10sttranscriptcluster.db") ??????????????

do nothing. this is to view the content/ functions of the package.

Symbols = unlist(mget(probes, hugene10sttranscriptclusterSYMBOL, ifnotfound=NA)) ???????????

This is to get the gene symbols. I don't know how to get for citrus

Entrez_IDs = unlist(mget(probes, hugene10sttranscriptclusterENTREZID, ifnotfound=NA)) ????????

This is to get the Entrez Ids. I don't know how to get for citrus

[zizigolu]

Thank you Deepak,

When I did normalization with my doupts I got a rma.txt like below, a long file (one column and many rows) of which I copied and pasted only few rows:

GSM1645665_120727-120222A_H_Citrus_.CEL
 1.7350
 1.7350
 6.5272
 6.7832
 6.9627
 7.8931
 7.8896
10.3757
 9.4220
11.4562
 8.4377
 9.7743
 8.3431
 8.2561
 9.3874
 7.8312
 8.7202
 5.5769
 5.5464
 5.7477
 6.1443
 5.4833
 6.2989
 7.7261
 5.9480
 6.6255
 6.8168
 6.8916

while the produced rma.txt by tutorial is like this:

#%guid=00003123-44eb-4068-1f22-004e560061c2
#%affymetrix-algorithm-param-apt-engine=ProbesetSummarizeEngine
#%affymetrix-algorithm-param-apt-program-name=Expression Console
#%affymetrix-algorithm-param-apt-command-line=
#%affymetrix-algorithm-param-apt-exec-guid=0000040472-1431761500-0000017680-0000004998-0000023182
#%affymetrix-algorithm-param-apt-analysis-guid=0000758a-0436-4e15-2043-004609003cbb
#%affymetrix-algorithm-param-apt-time-str=Sat May 16 12:01:40 2015
#%affymetrix-algorithm-param-apt-version=1.4.1.46
#%affymetrix-algorithm-param-apt-cvs-id=.........................................................

I think by my doubts I could not get the proper result