Hawaiian black coral (Antipatharia) complete mitochondrial genomes have limited phylogenetic signal for taxonomic resolution of species

Leah Shizuru1*, Van Wishingrad1*, Kenji Takata1,2, Anthony Montgomery3, Daniel Wagner4, Brian W. Bowen1, and Robert J. Toonen1#

1Hawai‘i Institute of Marine Biology, University of Hawai‘i at Mānoa, Kāne‘ohe, HI, United States of America
2Graduate School of Agricultural and Life Sciences, The University of Tokyo, Yayoi, Tokyo, Japan
3US Fish and Wildlife Service, Pacific Fish and Wildlife Office, Honolulu, HI, United States of America
4Ocean Exploration Trust, Honolulu, HI, United States of America

*These two authors contributed equally to this work

#Author for correspondence: toonen@hawaii.edu
 

SCRIPT FOR DE NOVO GENOME ASSEMBLY

#!/bin/bash
#Script to trim & filter raw reads and produce then evaluate a de novo genome assembly 
#Written by Evan W. Barba, modified by Leah E.K. Shizuru 
#This pipeline depends on:
#Trim Galore! http://www.bioinformatics.babraham.ac.uk/projects/trim_galore/
#SPAdes https://github.com/ablab/spades
#QUAST https://github.com/ablab/quast

###Begin Code###

if [ ! -d "trim_galore" ]; then
		mkdir trim_galore
			fi

#cd into the directory where fastq.gz files are located and employ the following to batch process on pair-end fastq.gz files
#files had the following naming scheme: "sampleID_raw_F.fastq.gz" and "sampleID_raw_R.fastq.gz"
#brace expansion enables Trim Galore! to grab all files that are named _raw_F.fastq.gz and _raw_R.fastq.gz and apply the specified trimming parameters
	
	find -name "*_raw_F.fastq.gz" | cut -d "_" -f1 | parallel trim_galore --illumina --paired --retain_unpaired --fastqc -o trim_galore/ {}\_raw_F.fastq.gz {}\_raw_R.fastq.gz
	
  	find ./ -name "*.F.fq.gz" | cut -d "." -f2 | sed 's/\///' > popslist

#SPAdes:

spades.py --careful -t 30 -m 300 -o /data/leah/spades/`sampleID` -1 `forward_trimmed_read` -2 `reverse_trimmed_read`

# QUAST:

	if [ ! -d "quast" ]; then 
		mkdir quast
			fi

	KMER=(` find ./ -maxdepth 1 -name "K*" | cat  `)
		ls K* -d | parallel -j 30 cp ./{}/final_contigs.fasta ./quast/{}.fasta
	
cd quast
	
quast.py -e -t 30 -m 300 ../contigs.fasta "${KMER[@]/%/.fasta}"

#create "large_contigs.fasta" 

	sed ':a;N;/^>/M!s/\n//;ta;P;D'  contigs.fasta > sed_corr.fasta
	
awk '/^>/ { getline seq } length(seq) >10000 { print $0 "\n" seq }' sed_corr.fasta > large_contigs.fasta

#BLAST large_contigs.fasta as a secondary check for assembly
 
IQ-TREE PARTITION COMMANDS

RaxML-style partition commands used for IQ-TREE analysis in Chapter 1

#NEXUS
begin sets;
	charset ATP6 = 1-717;
	charset ATP8 = 718-939;
	charset COX1 = 940-4295;
	charset COX2 = 4296-5048;
	charset COX3 = 5049-5837;
	charset CYTB = 5838-7025;
	charset ND1  = 7026-8102;
	charset ND2  = 8103-9677; 
	charset ND3  = 9678-10037; 
	charset ND4  = 10038-11540;
	charset ND4L = 11541-11840;
	charset ND5  = 11841-16153;
	charset ND6  = 16154-16822;
end;
 
RaxML-style partition commands used for IQ-TREE analysis 

#NEXUS
begin sets;
	charset ATP6 = 1-702;
	charset ATP8 = 703-915;
	charset COX1 = 916-3436;
	charset COX2 = 3437-4186;
	charset COX3 = 4187-4975;
	charset CYTB = 4976-6118;
	charset ND1  = 6119-7169;
	charset ND2  = 7170-8744; 
	charset ND3  = 8745-9128; 
	charset ND4  = 9129-10604;
	charset ND4L = 10605-10904;
	charset ND5  = 10905-14928;
	charset ND6  = 14929-15573;
end;
 
SCRIPT FOR IQ-TREE

#!/bin/bash
#Script used for IQ-Tree

Iqtree -s /`path to alignment in .fasta format` -p /`path to partitions.nex` -m MFP+MERGE -B 1000 -alrt 1000 -bnni -T auto