Dear Heidi, other users. I have an SDS formatted file to import using raw <- readCtData(files=c("SDS2.txt"), path=path, SDS=TRUE) Error in `[.data.frame`(sample, , Ct) : undefined columns selected This is done naively without setting arguments, which may well solve things. I will try it. However, the SDS file seems to have 3 parts to it. 1) The header, which is skipped when using SDS=TRUE, 2) the raw data in tab delimited (the plate ID is missing and as there are multiple plates, I am thinking I should add that in) and 3) There is a summary section at the end, quite long. Does readCtData understand the third summary data section? Or are the SDS files I have slightly strange? Also to extract raw data with Perl, I assume I need; 1) well number 2) plate id 3) sample 4) detector 5) task 6) CT Is this enough information to carry out all tests with HTqPCR? Thank you. 1) header section SDS 2.2.2RQ Results 1.0 Filename Card A 1-10.sdm Assay Type RQ Study EmbeddedFile Card A (1) BX005 BX010 BX012 BX013 A Run DateTime Fri May 22 11:53:04 BST 2009 Operator ThermalCycleParams EmbeddedFile Card A (2) BX007 BX008 BX011 Ref A Run DateTime Wed Jul 22 14:38:58 BST 2009 Operator .......etc, then 2) raw tab delim data Well PlateID Sample Detector Task Ct delta Rn delta Ct Ct Avg Ct SD Avg Delta Ct delta Ct SD Endo Ct Avg Endo Ct SD delta delta Ct RQ RQ Min RQ Max Baseline Type Baseline Start Baseline Stop Threshold Type Threshold Instrument Well Status Rejected Filename 1 BX005 CSF3-Hs00236884_m1 Target Undetermined 0.12634672 40.0 20.051275 19.948725 Manual 3 15 Manual 0.60596806 201488 Card A (1) BX005 BX010 BX012 BX013 A 2 BX005 CSF2-Hs00171266_m1 Target 35.817932 3.3153937 35.817932 15.869207 19.948725 0.0 1.0 Manual 3 15 Manual 0.7333056 201488 Card A (1) BX005 BX010 BX012 BX013 A ..........etc, then finally 3) summary data. Summary Data for Study Card A 1-10.sdm Endogenous Control 18S-Hs99999901_s1 Calibrator BX005 Multiplicity Non-Multiplex PlateID Sample Detector Avg delta Ct delta Ct SD delta delta Ct RQ RQ Min RQ Max Threshold Sample Status BX005 18S-Hs99999901_s1 0.59327465 BX010 18S-Hs99999901_s1 0.59327465 BX012 18S-Hs99999901_s1 0.59327465 BX013 18S-Hs99999901_s1 0.59327465 [[alternative HTML version deleted]]

Hi John > Dear Heidi, other users. > I have an SDS formatted file to import using > raw <- readCtData(files=c("SDS2.txt"), path=path, SDS=TRUE) > Error in `[.data.frame`(sample, , Ct) : undefined columns selected > I suspect this is because the SDS input in your file is a slightly different format than the other SDS files I've encountered. It's possibly an issue of SDS 2.2 versus 2.3? > This is done naively without setting arguments, which may well > solve things. > I will try it. Yep, that should hopefully solve it. Especially, readCtData per default tries to read in data from the following columns (/parameters): flag = 4, feature = 6, type = 7, position = 3, Ct = 8 This corresponds to a file where the headers are: # Plate Pos Flag Sample Detector Task Ct Delta Ct Avg Delta Ct ....etc.... Since your headers are apparently different/in a different order, you'll need to set these 5 parameters accordingly. qPCR data has a nasty habit of coming in slightly varying formats, so if it still doesn't work gimme a shout and I'll dig a bit deeper. > However, the SDS file seems to have 3 parts to it. 1) The > header, which is skipped when using SDS=TRUE, 2) the raw data in tab > delimited (the plate ID is missing and as there are multiple > plates, I am > thinking I should add that in) and 3) There is a summary section at > the end, > quite long. > > Does readCtData understand the third summary data section? Or are > the SDS > files I have slightly strange? > That part is actually skipped completely. What happens with SDS=TRUE is that the first 100 lines of the file are read in to determine the length of the header. This header is omitted, and the following n.features*n.data lines are read. Hence, if you say n.features=384 and n.data=1, only the first 384 lines of data will be read in, even if the files contains e.g. data from 5 PCR cards. If however you say n.data=3, then 1152 lines of data will be loaded, and formatted into a 384 x 3 matrix. > Also to extract raw data with Perl, I assume I need; > > 1) well number > 2) plate id > 3) sample > 4) detector > 5) task > 6) CT > > Is this enough information to carry out all tests with HTqPCR? The minimal amount of information needed is really just the Ct values themselves, and some sort of ID for each. Well number can be useful for checking e.g. spatial artifacts on the plate, but is not strictly required. Some sort of sample or plate ID is mainly for your own purpose; if missing they're per default named Sample1, Sample2 etc. HTH \Heidi > > Thank you. > > 1) header section > > SDS 2.2.2RQ Results 1.0 > Filename Card A 1-10.sdm > Assay Type RQ Study > EmbeddedFile Card A (1) BX005 BX010 BX012 BX013 A > Run DateTime Fri May 22 11:53:04 BST 2009 > Operator > ThermalCycleParams > EmbeddedFile Card A (2) BX007 BX008 BX011 Ref A > Run DateTime Wed Jul 22 14:38:58 BST 2009 > Operator > .......etc, then 2) raw tab delim data > > Well PlateID Sample Detector Task Ct delta Rn delta Ct Ct Avg Ct SD > Avg > Delta Ct delta Ct SD Endo Ct Avg Endo Ct SD delta delta Ct RQ RQ Min > RQ Max Baseline > Type Baseline Start Baseline Stop Threshold Type Threshold > Instrument Well > Status Rejected Filename > 1 BX005 CSF3-Hs00236884_m1 Target Undetermined 0.12634672 40.0 > 20.051275 > 19.948725 Manual 3 15 Manual 0.60596806 201488 Card A (1) BX005 > BX010 BX012 > BX013 A > 2 BX005 CSF2-Hs00171266_m1 Target 35.817932 3.3153937 35.817932 > 15.869207 > 19.948725 0.0 1.0 Manual 3 15 Manual 0.7333056 201488 Card A (1) > BX005 BX010 > BX012 BX013 A > > ..........etc, then finally 3) summary data. > > Summary Data for Study Card A 1-10.sdm > Endogenous Control 18S-Hs99999901_s1 > Calibrator BX005 > Multiplicity Non-Multiplex > PlateID Sample Detector Avg delta Ct delta Ct SD delta delta Ct RQ > RQ Min RQ > Max Threshold Sample Status > BX005 18S-Hs99999901_s1 0.59327465 > BX010 18S-Hs99999901_s1 0.59327465 > BX012 18S-Hs99999901_s1 0.59327465 > BX013 18S-Hs99999901_s1 0.59327465 > > [[alternative HTML version deleted]] > > _______________________________________________ > Bioconductor mailing list > Bioconductor@r-project.org > https://stat.ethz.ch/mailman/listinfo/bioconductor > Search the archives: http://news.gmane.org/ > gmane.science.biology.informatics.conductor [[alternative HTML version deleted]]