Search for a set of compatible indexes for your sequencing experiment according to:
- the number of samples
- the desired multiplexing rate (i.e. number of samples per pool/lane)
- the constraint on the indexes (none, use each one or each combination only once)
The list of the available indexes must be stored in a text file containing two tab-separated columns (without header): index ids are in the first column and the corresponding sequences in the second. inputIndexesExample.txt is an example of such a file and can be used to test checkMyIndex. For dual-indexing sequencing experiments the user needs to provide two files: the first one for the indexes 1 (i7) and the second for the indexes 2 (i5). index24-i7.txt and index24-i5.txt are available to test the research of compatible dual-indexes. Moreover, two files index96_UDI-i5.txt and index96_UDI-i7.txt are available to test the application in a Unique Dual-Indexing (UDI) context.
Click here to use the shiny interface of checkMyIndex.
If both shiny and shinyjs R packages are already installed, one can use the application locally running the two following lines in R:
library(shiny)
runGitHub("PF2-pasteur-fr/checkMyIndex", launch.browser=TRUE)
checkMyIndex can be executed calling checkMyIndex.r with Rscript. As checkMyIndex.r sources global.r, both files must be placed in the same directory. Here are 4 examples using either inputIndexesExample.txt or index24-i7.txt and index24-i5.txt:
- List of 9 indexes for 9 samples distributed on 3 lanes with the four-channel Illumina chemistry:
Rscript checkMyIndex.r --inputFile7=inputIndexesExample.txt -C 4 -n 9 -m 3
- List of 12 indexes for 12 samples distributed on 4 lanes with the four-channel Illumina chemistry using each lane combination only once:
Rscript checkMyIndex.r --inputFile7=inputIndexesExample.txt -C 4 -n 12 -m 3 -u lane
- List of 12 indexes for 12 samples distributed on 4 lanes with the two-channel Illumina chemistry using each index only once:
Rscript checkMyIndex.r --inputFile7=inputIndexesExample.txt -C 2 -n 12 -m 3 -u index
- List of 24 dual-indexes for 24 samples distributed on 2 lanes with the two-channel Illumina chemistry:
Rscript checkMyIndex.r --inputFile7=index24-i7.txt --inputFile5=index24-i5.txt -C 2 -n 24 -m 12
The help page of the script can be displayed running the following command:
Rscript checkMyIndex.r --help
Here is the list of the R packages needed to run checkMyIndex:
-
shiny and shinyjs to run the shiny application locally
-
optparse to interpret the input parameters when using the Rscript command line
-
parallel to speed up the calculations
One can install each of these packages running install.packages(packageName) in R.
Illumina has developed three types of chemistry: the four-channels for the HiSeq and MiSeq sequencing devices, the two-channels for the NovaSeq, NextSeq and MiniSeq devices and the one-channel for the iSeq 100 device. With the four-channel chemistry, a red laser detects A/C bases and a green laser detects G/T bases and the indexes are compatible if there is at least one red light and one green light at each position. With the two-channel chemistry, G bases have no color, A bases are orange, C bases are red and T bases are green and indexes are compatible if there is at least one color at each position. Note that indexes starting with GG are not compatible with the two-channel chemistry. With the one-channel chemistry, compatibility cannot be defined with colors and indexes are compatible if there is at least one A or C or T base at each position. Please refer to the Illumina documentation for more detailed information on the different chemistries.
- Version 1.0.1: update of the user interface with shinyjs
- Version 1.0.2: deal with the new Illumina unique dual-indexes (UDI)
This tool has been developed at the Biomics pole of the Institut Pasteur by Hugo Varet ([email protected]) and an Application Note describing it has been published in 2018 in Bioinformatics. Please note that checkMyIndex is provided without any guarantees as to its accuracy.