1. Background

Now that we have filtered and formatted LepMap files, we can start the linkage mapping stage of this project. The workflow for LepMap is as follows:

Parent Call > Separate Chromosomes > Join Singles to All > Order Markers <

DISCLAIMER: the paths in this were simplified from my original analyses, which ran on the JCU HPC. They might not work as is if you try and replicate these analyses. You will have to adjust them accordingly.

2. ParentCall2

#!/bin/bash

#PBS -N ustacks-array
#PBS -l ncpus=1
#PBS -l mem=5g

##### Change Directory #####
cd ~/ProjectDirectory/2_LinMap

#### Run parent call #####
java -cp 0_lepmap/bin/ ParentCall2 data=1_DataPrep/5_2_LepMapInput_HC_A.txt > 6_1_parcall_HC

Submit with

qsub 6_1_ParentCall.sh

3. SeparateChromosomes2

I do this both with informativeMask=1 and informativeMask=2, but separately for each.

#!/bin/bash

#PBS -N SepChrom2    
#PBS -l ncpus=1
#PBS -l mem=10g

##### Change Directory #####
cd ~/ProjectDirectory/2_LinMap

#### Run parent call #####
java -cp 0_lepmap/bin/ SeparateChromosomes2 data=6_1_parcall_HC lodLimit=40 sizeLimit=20 > 6_2_sepchrom_HC
qsub 0_scripts/2_2_SeparateChromosomes2.sh &> 0_logfiles/2_2_separateChromosomes2.log

Before moving forward, let’s check the size of linkage groups

cut -f 1 ../2_LinMap/6_2_sepchrom_HC | sort | uniq -c | tail -n 14
##  153 0
## 1101 1
##  440 10
##  345 11
##  334 12
##  302 13
## 1088 2
## 1057 3
##  941 4
##  748 5
##  729 6
##  535 7
##  468 8
##  461 9

4. JoinSingles2All

#!/bin/bash

#PBS -N joinsingles2all
#PBS -l ncpus=1
#PBS -l mem=10g

##### Change Directory #####
cd ~/ProjectDirectory/2_LinMap

#### Run parent call #####
java -cp 0_lepmap/bin/ JoinSingles2All data=6_1_parcall_HC map=6_2_sepchrom_HC lodLimit=10 iterate=1 > 6_3_joinsingles_HC

Submit with qsub as usual.

Now let’s check the LG size after this step

cut -f 1 ../2_LinMap/6_3_joinsingles_HC | sort | uniq -c | tail -n 14
##   21 0
## 1118 1
##  447 10
##  360 11
##  340 12
##  307 13
## 1095 2
## 1075 3
##  945 4
##  772 5
##  741 6
##  542 7
##  475 8
##  464 9

Done! These steps are pretty straight forward. The most work usually goes into identifying the right number of linkage groups. For this project we already knew that most Litoria species have 13 chromosomes, and because of the large sample size we identify the linkage groups easily.