1. A Metagenome Letdown
By Rebekah McCurdy

2. Claiming A Sequence OctHS.APNO1912-b2 OctHS.APNO1912-g2*****
GTTAATTCGCGCAATTCTGCCATTACGTCATGCAACTCAGGAGCGTCATAACGCGCCGTCTCAAAACTTTTTTTATCAAATTCTATAATTTGCGTTATTCGTTGTTGTGTTTAATATCAATATCACCATACATTATGAATAAGCTATATCTAGATAAAAATGCCTTAACCGCATTGCCTCTAATCACCATGTCTAAGTCGTGGAGTTCGATAGCATAATCGCCACGTTCTATATAGTTGCGGAATATCTCTATTGCGTCTCTAGCTGTTTTTAGTAGTTGTTCTTTTTGTGTTAGGGTTTTTATTTCGGTTTTTGTTGCCATGCCAGTAAGAAAACTGTGGATTTAAAAGCTTTTTGTTTATAATATGTCAACCTCATTCTTTACTTGTATCTAATGCATGTCGGGGTACATATTTTATATTCCGCTTAATGTGTTTATGTTAGTAAAGGGCCACGTTTACACAAAAACACGGCGCAGGAAAAAGGCGCGCAATGCGCGTCTCCACACTTTGTAAACCTTACCCTTTACATAGAAATTTCTAAAGCCAAATATCTAGCGATATGTGTCAAACATAGGATTTACACTTATCCAACGCTACATTTTACATGTGTCAAGAGACTGAAAACATGCGCCACGCATGCCGCACATTATCTCTTTTCACAGATTTTTTCAACGGCGCCAATATATCGATATATATCTAATCCCATACTTTACCCATGTCCAGCAGGACCCCCCAAAAAATTTCCAATTTTTCCTTTAAAATCCAAAATTTTTTTCCTTTAAAAAAATGGAATATTTCCCAAAAAAAGGGGGGGCCCCCCCCGGGGGCCCCCTCCCCTTTACAAAAAGCCTTTTTCCAAAAGGGGGCAACCCCGGGT
OctHS.APNO1912-b2
OctHS.APNO1912-g2*****

3. Blast Results
Seq NGACTGAGACTGCGGC-GCGA-TTCGGATCCATTGATGACC
Seq NNGGTCTGAGACTGCGGC-GCGA-TTCGGATC-ATTGATGACC
Seq NGGGTCTGAGACTGCGGC-GCGA-TTCGGATC-ATTGATGAC
Seq NGACTGAGACTGCGGC-GCGA-TTCGGATC-CTTGATGACT
Seq NGGTCTGAGACTGCGGCCGCGA-TTCGGATCCATTGATGAC
Seq NNNGGTCTGAGACTGCGGC-GCGA-TTCGGATCCATTGATGACA
Seq NGGACCGAGACTGCGGCCGCGA-TTCGGATCCATTGATGACC
Seq ACTGAGACTGCGGC-GCGA-TTCGGATCCATTGATGAC
Seq CTCAGACTGCGGC-GCNA-TTCGGATCC-TTGATGAA
Seq NGTCTGAGACTGCGGC-GCGA-TTCGGATCCATTGATGACC
Seq NGTCTGAGACTGCGGC-GCGA-TTCGGATC-ATTGATGAC
Seq NNNGGTCTGAGACTGCGGC-GCGA-ATTGGATCCATTGATGACA
Seq NNGGTCTGAGACTGCGGCCGCGA-TTCGGATCCATTGATGAC
Random contigs from Octopus, similar beginning sequence...what is it?

4. Editing The Sequence Linker sequences!! N-nucleotides-unknown
NGTCTGAGAC TGCGGCGCGA TTCGGATCCT TGATGACGAT ACAAGCGGCC GCATC
(Specific to my read)

5. Extending the Sequence
Significant Overlaps
OctHSe.APNO1912-g2(my read) nucleotides match
OctHSe.ATYB2970-b2 from nucleotides
E-value: 2.0d-140 (good low number)
% Identity:84.64
Length is 940, last five nucleotides are insignificant (similar enough)
Extends the left side by adding to beginning of original read

6. Repeat Process Keep going!! Extend Extend!!
Got 2 more clean overlaps before stopping due to lack of clean overlaps
Length now 1374 nt long (was 880nt)

7. Extension on only One end?
Initial Blast results up to coordinate 727
But why?
Blasting coordinates results in one small match
AAAAAAGGGG GGGCCCCCCC C (20 nt) small match in middle of sequence no significance found
No clean overlaps to extend.

8. GeneMark Results 3 Predicted proteins: >gene_1
IVQDEILGTIRQVLTIMPNEISLRKWFDRHYGETTLRRIRLLKPYKGVDTIIELVRGFEG EGNNWWRSVDIVPRDTNHITALLSKIRDFDTFTNTLGCIFKVCRDMSYVNVVECDISACL
>gene_2
MATKTEIKTLTQKEQLLKTARDAIEIFRNYIERGDYAIELHDLDMVIRGNAVKAFLSRYS LFIMYGDIDIKHNNEITQIIEFDKKSFETARYDAPELHDVMAELRELTVEKVKEKVHKIL SKIQ
>gene_3
PGVAPFWKKAFCKGEGAPGGGPPFFWEIFHFFKGKKFWILKEKLEIFWGVLLDMGKVWD

9. Blasting the Proteins 1st Predicted protein results:
No significant results found
E-values all above 1.0
Score E Sequences producing significant alignments: Score (Bits) E-Value
ref|YP_ | diguanylate cyclase [Nitrobacter winogradsky ref|YP_ | valyl-tRNA synthetase [Natranaerobius the ref|YP_ | LuxR transcriptional regulator [Ralstonia eu ref|NP_ | chemotaxis regulator CheZ [Ralstonia solanac

10. 2nd Predicted Protein results: -No significant results found
-e-values all above 2.0
Score E Sequences producing significant alignments: Score (Bits) E-Value
ref|XP_ | hypothetical protein TRIADDRAFT_ ref|NP_ | protein BRE [Zea mays] >gb|ACF ref|YP_ | nitrite/sulfite reductase hemoprotein beta s ref|NP_ | ATPase, Class II, type 9A [Sus scrofa] ref|ZP_ | hypothetical protein CIT292_ gb|EEH | oxidoreductase [Citrobacter sp. 30_2]

11. 3rd Predicted Protein results:
No significant results found
e-value (one result) is 5.7
Sequences producing significant alignments: Score(Bits) E-Value
ref|XP_ | PREDICTED: similar to DNA repair/transcription protein

12. Phylogenetic Tree
When trying to get sequences similar to mine in all known sequences, all results were ‘nil’ so I have no tree to display.

13. Conclusion Significant lack of similar proteins
Leads to no phylogenetic tree
Extension on only one side
3 predicted proteins-no promising results