---
title: 'LIFE_pilot_step4_colony2'
disqus: hackmd
---

LIFE pilot step 3: confirming pedigree reconstructions using Colony2
===
by Tanya Lama

Contact me at [e-mail] re: questions about this method

## Objective
pedigree reconstruction from SNP data using Colony2

## Dependencies
Input is the threshold 1.5 genotyping dataset described in step 2. The inputs are the same as those for Cervus. See the /folder for examples. Mind the absence of headers.

## Table of Contents

[TOC]

## Step 1: installing dependencies
### Install Colony2
Colony2 can be downloaded for PC/MacOS/Linus [here](https://www.zsl.org/node/40009/done?sid=191392&token=7a8113ab1ddf1fade37583372c5408a4). Note that for our volume of data we will be using the Linux version on the cluster whenever possible. The details here show how Colony2 can be installed on the cluster.

1. Download the above zip folder for linux to your desktop
2. In the terminal, use the scp command to send the folder from your desktop to your /bin folder on the cluster

scp /Users/tanya/Desktop/Colony2_Lnx_25_07_2021.zip [e-mail]:/home/tl50a/bin 

3. Unzip the folder into a colony2 subfolder in /bin
4. For a serial run of COLONY we will use colony2s.ifort.out. Make it executable using

chmod +x colony2s.ifort.out

5. COLONY will be executed in R using the run.colony function. However, it can also be called directly using the below command. Give it a try to be sure the program runs properly on the test dataset.

./colony2s.ifort.out IFN:YourInputFileName

## Step 2: preparing inputs for build.colony.input

## Step 3: creating a *.DAT input for Colony2 using build.colony.input in R

Move your *.DAT input from your desktop to the cluster so that it can be run with Colony2

scp /Users/tanya/Desktop/Colony2.DAT [e-mail]:/home/tl50a/colony/data

## Step 3: run Colony2
1. Request an interactive session on the cluster so as not to run Colony on the head node

bsub -q interactive -W 120 -Is bash
module load R/4.0.4_gcc
module load gcc/8.1.0

#Enter an R session with R

run.colony(colonyexecpath="prompt",datfilepath="prompt",wait=FALSE,monitor=TRUE)

bin:
/project/uma_lisa_komoroske/bin/colony2/colony2s.ifort.out IFN:portugal_lynx
input:
/home/tl50a/colony/data/

colony2.dat

Cohort 2:
2 offspring
6 mothers
4 fathers

Cohort 3:
8 offspring
9 mothers
8 fathers

Cohort 4:
11 mothers
18 Offspring
9 fathers

Edit your c4 inputs (mothers, fathers, offspring)
Copy/paste into portugal_cohort 4.dat
Use the following command to transfer them to the cluster
scp

scp /Users/tanya/Desktop/portugal_cohort4.dat [e-mail]:/home/tl50a/colony/data/colony2.dat

## Appendix and FAQ

## Notes from Edoardo Velli re: non-invasive genetic monitoring with scats
scat collection & robot?
filter or bead DNA extraction --> filter. They use QUIAGEN QCUBE for 12-sample automated extraction. Risk of contamination is extremely low, which is important to fecal samples.

Plate extractiong (96) on another automated robot. Risk of contamination may be higher, but Edoardo has not had that issue. As long as you include controls to confirm. The most important thing is that the samples are well-filtered so. that the samples don't clog things up. Filter-based extraction has not been our first choice because materials in the fecal samples are clogging the filter. Using the swab we can avoid this. 10 minute or more of frotar so that solids go down and desirable materials float to the top. It takes about an hour for 6 samples.

blood & tissue quiagen kit. they have tried the stool kit but it wasn't better than the blood & tissue kit

fecals -- 2 types of samples. 1 is a swab in an ATL buffer that comes with the quiagen kit. these samples preserve themselves very well. If the fecal is dry, you can use water to reconstitute

swabs -- plastic and cotton.
swab the top and the sides, not the bottom because

4 replicas y si tienen algun problema hacen unos 4 replicas mas
2-4 controls in the center and on the sides

:::info
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###### tags: `iberian_lynx` `fluidigm` `genotyping` `SNP`