1. M. roreri de novo genome assembly using abyss/1.9.0-maxk96
Abyss 1.9.0: introduces a new tool called Sealer for closing scaffold gaps. Also, it has Konnector, a fast and memory-efficient tool to fill the gap between paired-end reads.
GROUP 5
Hyeim Jung
Pedro Pablo Parra
Diana Vanessa Sarria Zuniga
Jacob Shoemake

2. Construction of contigs Solving Ambiguities and merging contigs
without using the paired-end information
Solving Ambiguities and merging contigs
Using paired-end information 1 2
HOW ABySS WORKS…
Assembly algorithm: two major steps
Required
Select a ABySS compiled version depending on a maximum k-mer size
K-mer size: Kmergenie
Input library files
Paired-end
Unpaired(Single-end)
Mate pair
The assembly is performed in two major steps.
First, without using the paired-end information, contigs are extended until either they cannot be unambiguously extended or come to a blunt end due to a lack of coverage.
In the second step the paired-end information is used to resolve ambiguities and merge contigs.
The paired-end information is used to identify contigs that can be linked together.
Two contigs are considered to be linked if at least p pairs (by default p = 5) join the contigs
Contain
Konnector: to fill the gap between paired-end reads
Sealer: for closing scaffold gaps

3. OUR ASSEMBLY STRATEGIES… Two assembly types
abyss-pe k=87 name=assembly5
lib='pe1' mp='mp1'
pe1=‘paired PE.1.fq paired PE2.fq’
se=’unpaired PE-MP’
mp1=‘paired MP.1.fq paired MP.2.fq’
Assembly 3
abyss-pe k=81 name=assembly3
lib='pe1 pe2' mp='mp1'
pe1=‘paired PE.1.fq paired PE2.fq’
pe2=‘paired MP.1.fq paired MP.2.fq’
se=’unpaired PE-MP’
mp1=‘paired MP.1.fq paired MP.2.fq’
Paired PE and Unpaired PE-MP 87
Paired PE-MP and Unpaired PE-MP 81
Note: mp1 is used for scaffolding. Do not contribute to the consensus sequence.

4. Assembly 3 Assembly 5 Contigs Contigs Scaffolds Scaffolds Paired MP
Paired PE
Paired PE
Paired MP
Unpaired
PE&MP
Unpaired
PE&MP
Scaffolds
Scaffolds
Paired MP
Paired MP

5. Evaluation of best assemblies
Quast Report without reference genome
Bowtie2
Assembly
File
# contigs
Largest
Total Length
N50
# N's
Predicted genes
Mapped PE reads
assembly_5
contigs.fa
(total, --min-contig 500bp) 4328
(>= 0 bp) 9711 (>= 1000 bp) 3544 (>= 5000 bp) 1887 (>= bp) 1181 (>= bp) 604 (>= bp) 268
553,471
(total, --min-contig 500bp) 57.68Mb (>= 0 bp) 58.59Mb (>= 1000 bp) 57.12Mb (>= 5000 bp) 52.99Mb (>= bp) 47.96Mb
(>= bp) 38.75Mb (>= bp) 27.02Mb
45,432
46,124
(unique) 17734
(>= 0 bp)
(>= 300 bp) 21553
(>= 1500 bp) 1189
(>= 3000 bp) 6
60.40% aligned concordantly exactly 1 time
22.51% aligned concordantly >1 times
Total 82.91%
scaffolds.fa
(total, --min-contig 500bp) (>= 0 bp) (>= 1000 bp) (>= 5000 bp) (>= bp) (>= bp) (>= bp)
1,036, ,564
(total, --min-contig 500bp) Mb (>= 0 bp) Mb (>= 1000 bp) 57.37Mb Mb (>= 5000 bp) Mb (>= bp) Mb (>= bp) Mb Mb (>= bp) Mb
99,290 51,001
568,877
945
(unique)
(>= 0 bp)
(>= 300 bp)
(>= 1500 bp)
(>= 3000 bp) 66 66
60.41% aligned concordantly exactly 1 time
22.54% aligned concordantly >1 times
Total % 
assembly_3
(total, --min-contig 500bp) 4816 (>= 0 bp) (>= 1000 bp) 3514 (>= 5000 bp) 1642 (>= bp) 1078 (>= bp) 567 (>= bp) 256
1,035,772
(total, --min-contig 500bp) 56.36Mb (>= 0 bp) 61.10Mb (>= 1000 bp) 55.45Mb (>= 5000 bp) 50.87Mb
(>= bp) 46.79Mb
(>= bp) 38.70Mb (>= bp) 27.77Mb
48,947
247,454
(unique) 17570
(>= 0 bp)
(>= 300 bp) 21274
(>= 1500 bp) 1171
(>= 3000 bp) 63
58.95% aligned concordantly exactly 1 time
22.55% aligned concordantly >1 times
Total 81.5%
(total, --min-contig 500bp) (>= 0 bp) (>= 1000 bp) (>= 5000 bp) (>= bp) (>= bp) (>= bp)
1,771, ,868
(total, --min-contig 500bp) Mb (>= 0 bp) Mb (>= 1000 bp) Mb (>= 5000 bp) (>= bp) Mb (>= bp) Mb (>= bp) Mb
102,079 51,480
1,600,849
806
(unique)
(>= 0 bp)
(>= 300 bp)
(>= 1500 bp)
(>= 3000 bp) 63 63
58.97% aligned concordantly exactly 1 time
22.62% aligned concordantly >1 times
Total 81.59% 
Evaluation of best assemblies
PE: , peak 301
MP: , peak 1700
Quast options: quast/3.2 --gene-finding --eukaryote
Bowtie2 options: bowtie2/ very-sensitive-local --no-unal --phred33 -p 20

6. conclusions Total Length of Assembly # Scaffolds Largest scaffold N50
Abyss assembly
Broken
Comment
Total Length of Assembly
(~)
Assembly 5
Assemblies: Same Broken: Assembly 3 has 1.1 Mb less.
# Scaffolds
Assembly 3 has many Scaffolds <500 bp compared with Assembly 5.
Largest scaffold
Assembly 3
N50
Assembly 3 (~)
Abyss: Assemb. 3 has 2,789 bp more Broken: Assemb. 3 has 479 bp more.
# N's
Abyss: Assemb. 3 has 1 Mb more N's. Broken: Assemb. 5 has 139 more N's.
# Unique predicted genes
Assembly 5 (~)
Abyss: Assemb. 5 has 67 genes more Broken: Assemb. 3 has 71 genes more
Mapped paired end reads
Assemb. 5 has 1.36% more (82.95% vs 81.59%).

8. reads; of these:
(100.00%) were paired; of these:
(17.09%) aligned concordantly 0 times
(60.40%) aligned concordantly exactly 1 time
(22.51%) aligned concordantly >1 times
----
pairs aligned concordantly 0 times; of these:
(61.27%) aligned discordantly 1 time
pairs aligned 0 times concordantly or discordantly; of these:
mates make up the pairs; of these:
37310 (1.11%) aligned 0 times
(21.66%) aligned exactly 1 time
(77.23%) aligned >1 times
99.93% overall alignment rate