1. Professors: Dr. Gribskov and Dr. Weil
AGRY-600 Genomics
Genome Assembly
Professors: Dr. Gribskov and Dr. Weil
Group 3:
Brett Lane Amanpreet Kaur
Stefanie Griebel Yulu Chen
Rupesh Gaire Akanksha Singh

2. Cleaned Data as Input Files
Genome Assembly Pipeline
Data Cleaning
REAPR
QUAST Gene finding
GapFiller
SSPACE
SPAdes
Kmergenie Kmer size
Cleaned Data as Input Files
Assembly
Group 3

3. Results Cleaning Steps - MP reads
Group 3

4. Kmergenie
Kmergenie was used to determine the size of k-mer for assembly.
All the reads – paired end and mate paired were used to predict k-mer size
Best kmer size predicted: 87
Predicted assembly size: 56.6 Mb
Group 3

5. SPAdes Genome Assembler - Why?
SPAdes is suitable for:
Illumina reads
Bacterial and fungal data
Small genomes not large genomes
Paired-end reads, Mate-pair reads and unpaired reads
Group 3

6. Assembly Statistics Group 3
N50
Longest
Contigs/ Scaffolds
Total Length
Program
Data
Comments
70,639 (Contigs)
74,736 (Scaffolds)
608,380
3293(Contigs)
3187(Scaffolds)
55.9 Mb
SPAdes
used multiple k-mer values: 51,61,71,81,83,87
PE, MP, both unpaired
#N's=2873
5.14 Ns/100 kbp
SPAdes allows De Bruijn graph assembly at multiple k-mer sizes, not just a single fixed one.
Merges different k-mer assemblies
Group 3

7. Scaffolding using SSPACE v3.0
The scaffolding was done using SSPACE v 3.0 using the Mate Pair Reads
Programme
Scaffolds
N50
Longest contig
Total length
#N’s per 100 kb
SPAdes
8073
(>=500bp=3187)
74.7 Kb
608 Kb
55.9 Mb
5.14
SSPACE v 3.0
5183
(>=500bp=967)
193 Kb
1.56 Mb
61.6 Mb
Group 3

8. Bridging the gaps using GapFiller
No. of N’s was very high after scaffolding :
#N’s per 100 Kb:
GapFiller was used for filling the gaps using Mate Pair reads
It reduced the number of N’s: /100 Kb
Scaffolds
N50
Longest contig
Total length
#N’s per 100 kb
Before gap filling
5183
(>=500bp=967)
193Kb
1.56 Mb
61.6 Mb
After gap filling
GapFiller highly reduced the gaps
Group 3

9. Mapping the PE reads to the Assembly
We used bowtie 2 to map the Paired end reads to our final assembly
59.58 % aligned concordantly exactly 1 time
24.01% aligned concordantly >1 times
Total: %
Overall alignment rate : %
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10. REAPR Error free bases: 85.75% Total Number of errors: 2652
FCD errors within a contig: 615
FCD errors over a gap: 46
Low fragment coverage within a contig: 146
Low fragment coverage over a gap: 1845
85 % of the bases are error free which is good
Group 3

11. Gene Prediction Genes were predicted using Quast
No. of predicted genes
Unique
17455
(>= 0 bp)
107219
(>= 300 bp)
22348
(>= 1500 bp)
1241
(>= 3000 bp) 77
Group 3

12. Conclusion Good assembly N50: 193Kb Longest contig : 1.56 Mb
Less no. of gaps
85.75 % bases are error free
Only concern is the total length of the genome (61 Mb)
Group 3

13. Thanks