RomainFeron / Nanopore_comparison

Comparison of several assembly and polishing methods for assembly of Nanopore and Illumina reads

Nanopore assembly method comparison

This repository contains the results of assembly quality assessment for several assembly and polishing methods for Nanopore and Illumina reads. The data and final genome assembly are included in Pan, Q.et al., (2020). The rise and fall of the ancient northern pike master sex determining gene. bioRxiv..

Information in this repository is not complete and will be updated soon.

Results

The full summary table including all assembly metrics and BUSCO metrics is available here. A short table with only the most important metrics is provided below:

Assembler	Reads	Racon rounds	Polca rounds	Pilon rounds	Scaffolds	Total scaffold size	Longest scaffold	Median scaffold size	Scaffold N50	Scaffold L50	Complete BUSCOs (%)	Single-copy BUSCOs (%)	Duplicated BUSCOs (%)	Fragmented BUSCOs (%)
canu	raw	0	0	0	3227	951948846	10942431	47627	1534791	149	43.7	42.9	0.8	7.4
canu	raw	1	0	0	3071	965393679	11078614	52248	1538696	150	56.7	55.5	1.2	7.9
canu	raw	2	0	0	3037	965700424	11087656	52831	1555114	149	57.6	56.6	1	8
canu	raw	2	0	1	3037	967423163	11110507	52883	1559591	149	94.5	90.6	3.9	2.8
canu	raw	2	1	0	3037	966320640	11094938	52855	1556531	149	74.1	71.7	2.4	9.1
flye	corrected	0	0	0	1329	916399443	29664458	100741	3800379	51	60	58.8	1.2	7.8
flye	corrected	1	0	0	1244	918414721	29727285	123939	3807314	51	59.2	57.7	1.5	8.1
flye	corrected	2	0	0	1231	918733717	29733036	127128	3807983	51	58.9	57.6	1.3	7.9
flye	corrected	2	0	1	1231	919922954	29788268	127613	3816572	51	94.4	90.9	3.5	2.8
flye	corrected	2	1	0	1231	918585399	29732637	127163	3808094	51	71.3	69.3	2	9.1
flye	raw	0	0	0	1757	947335101	35593798	10348	10807385	25	60	58.7	1.3	8.3
flye	raw	1	0	0	1401	942065221	35433054	21075	10751775	25	59.9	58.6	1.3	8.2
flye	raw	2	0	0	1352	941676606	35430667	23319	10747330	25	59.8	58.3	1.5	8.6
flye	raw	2	0	1	1352	942304953	35434112	23311	10752412	25	94.5	90.9	3.6	2.9
flye	raw	2	1	0	1352	941422740	35421523	23322	10749561	25	72.1	70	2.1	8.9
flye	trimmed	0	0	0	1699	908968079	17279927	133056	1983562	100	59.9	58.7	1.2	7.8
flye	trimmed	1	0	0	1648	911198514	17324657	142987	1991917	100	59.7	58.2	1.5	7.4
flye	trimmed	2	0	0	1634	911564900	17331222	145624	1993190	100	59.2	57.7	1.5	7.4
flye	trimmed	2	0	1	1634	913040417	17363090	145919	1990260	100	93.9	90.4	3.5	3
flye	trimmed	2	1	0	1634	911410720	17326448	145629	1989349	100	71.6	69.5	2.1	8.5
wtdbg2	corrected	0	0	0	2810	885831968	23252592	23709	3525371	60	43	42.7	0.3	6.3
wtdbg2	corrected	1	0	0	2597	895437456	23513364	26764	3559418	60	56.5	55.3	1.2	8.1
wtdbg2	corrected	2	0	0	2535	895312360	23529299	27517	3560626	60	56.9	55.7	1.2	7.9
wtdbg2	corrected	2	0	1	2535	896816104	23571653	27471	3562908	60	93	89.5	3.5	2.7
wtdbg2	corrected	2	1	0	2535	895293217	23530634	27476	3560501	60	70.4	68.5	1.9	8.5
wtdbg2	raw	0	0	0	1227	867381815	27191330	22375	7356404	32	33.4	33	0.4	5.7
wtdbg2	raw	1	0	0	1132	879206707	27550322	26886	7458027	32	56.2	54.8	1.4	8.1
wtdbg2	raw	2	0	0	1120	879589208	27562967	27355	7460231	32	57.2	56.2	1	7.9
wtdbg2	raw	2	0	1	1120	880869069	27593451	27398	7473325	32	93.2	89.8	3.4	2.5
wtdbg2	raw	2	1	0	1120	879576357	27563414	27350	7462154	32	69.6	67.9	1.7	8.3
wtdbg2	trimmed	0	0	0	2792	877426181	17149271	29942	2183325	94	43.4	42.8	0.6	6.9
wtdbg2	trimmed	1	0	0	2661	886838528	17322380	34351	2209111	94	56.9	55.8	1.1	7.7
wtdbg2	trimmed	2	0	0	2613	887016948	17333162	35749	2211406	94	57.8	56.8	1	7.8
wtdbg2	trimmed	2	0	1	2613	888765491	17357701	35797	2219808	94	93.7	90.2	3.5	2.7
wtdbg2	trimmed	2	1	0	2613	886978594	17330674	35728	2211370	94	70.4	68.3	2.1	8.8

Reads: raw means original Nanopore reads, corrected means Nanopore reads corrected by Canu, trimmed means Nanopore reads corrected and trimmed by Canu

Data

Nanopore reads

Nanopore reads were obtained from two individuals (two rounds of sequencing). Methods for the first round are described in Pan et al., 2019, in the Methods section Genome sequencing and assembly. Methods for the second round are described in Pan et al., 2020, in the Methods section Genome and population genomics sequencing.

TODO: stats for all reads

Illumina reads

Methods

This should have been a snakemake workflow but it was not. Next time... Instead examples of the commands used for each software are provided below.

Assembly

Canu

Flye

wtdbg2

Polishing

Racon

Pilon

ASSEMBLY=<assembly_file>

bwa index $ASSEMBLY

bwa mem -t 24 $ASSEMBLY \
<reads_file_1> \
<reads_file_2> | \
samtools view -bh > $ASSEMBLY.bam

samtools sort -@ 24 -o $ASSEMBLY.sorted.bam $ASSEMBLY.bam

samtools index $ASSEMBLY.sorted.bam

pilon -Xmx230G \
--genome $ASSEMBLY \
--frags $ASSEMBLY.sorted.bam \
--output <output_file> \
--changes \
--fix all \
--threads 24

Polca

polca.sh \
-a <assembly_file> \
-r '<reads_file_1> <reads_file_2>' \
-t 32