Genomika: Informácie ku trackom

Informácie k predmetu Genomika

Na tejto stránke sú informácie k trackom ktoré budete vytvárať na browseri (obe skupiny). K niektorým trackom pridáme ďalšie informácie v nasledujúcich dňoch.

Comments to the task list

• Task (A) is a prerequisite of all other tasks, the rest are mostly independent of each other.
• Tasks are marked as fast (no significant computation required), medium (estimated computation up to 1 hour), slow (longer computation, possibly several hours).
  • These times are only estimates, reality may vary. Perhaps provide actual running times (approximate) in your documentation.
  • Fast tasks can be done entirely on genomika server.
  • Students having accounts on compbio research cluster may run medium and slow tasks there.
• If you get stuck on one task, you can try to do at least initial stages of another one. Coordinate within group!
• Document your work. Documentation should be independent of this page and of the documentation created last year - copy and modify relevant passages, cite sources.

Basic information on creating tracks

• https://github.com/fmfi-genomika/genomika-2017/wiki/Basics-of-creating-tracks
• https://github.com/fmfi-genomika/genomika-2017/wiki/How-to-add-track-to-DB-and-display-it-on-page
• Important: compared to last year, path /kentsrc/kent/src/hg/makeDb/trackDb/ was moved to /kentsrc/trackDb/

(A) Genome (fast)

• Download genome in fasta format, add to browser
• Data malGlo [1], malSym [2]
• https://github.com/fmfi-genomika/genomika-2017/wiki/Setting-up-a-Yarrowia-lipolytica
• Important step not described is to rename chromosomes/contigs to something reasonable
• Genome versions are numbered, we will start with malGlo1 and malSym1

hgsql hgcentral -e '
insert into dbDb values (...);

insert into defaultDb values ("M. ingens","magIngA4");

insert into genomeClade values ("M. ingens","other",10);

(B) Protein coding genes and other items from the annotation (fast, needs A)

• Download genome annotation in GFF format, process to genepred format, split into two tracks: genes and other items
• Last year done by 2 groups based on 2 different databases:
• https://github.com/fmfi-genomika/genomika-2017/wiki/Protein-coding-genes-from-NCBI-RefSeq
• https://github.com/fmfi-genomika/genomika-2017/wiki/Protein-coding-genes-from-EnsembleFungi
• Coordinate with renaming chromosomes in step (1)
• Use appropriate IDs for naming genes
• Last-year tracks not ideal, try to improve
  • mRNA items in other item track are redundant, should be omitted
  • also items covering entire chromosome (type=region) should be omitted
  • protein coding genes could be displayed with codon highlighting - use the following settings in trackDb.ra:

baseColorUseCds given
baseColorDefault genomicCodons

(C) RepeatMasker (slow, needs A)

• Run RepeatMasker program to find repeated sequences (use fungi as species)
• https://github.com/fmfi-genomika/genomika-2017/wiki/RepeatMasker
• RepeatMasker and repeat library installed at compbio and genomika servers, or request a copy for your computer, registration may take several days

(D) tRNAscan-SE (medium, needs A)

• Run software for finding tRNA genes (for comparison with annotation)
• Download software from http://lowelab.ucsc.edu/tRNAscan-SE/ (already installed on compbio servers as tRNAscan-SE command)
• Convert output by script rna/tRNAscan-SEtoBED.py on github
• trackDb.ra record:

track tRNAs
shortLabel tRNA Genes
longLabel Transfer RNA Genes Identified with tRNAscan-SE
group genes
visibility hide
color 0,20,150
type bed 12
nextItemButton on
priority 10

(E) Augustus (slow, needs A)

• Run gene finder Augustus, create track with predicted genes (for comparison with annotation)
• Download and install software from http://bioinf.uni-greifswald.de/augustus/
  • Already installed on compbio servers
• Example of command line: augustus --uniqueGeneId=true --species=ustilago_maydis genome.fa > augustus.gtf
• ustilago_maydis is a related fungal species used for training parameters
• The result needs to be converted from gtf to genepred, by gtfToGenePred (at genomika server) with option -genePredExt
• If you name your track augustus, genome browser will recognize it automatically, no need to modify trackDb.ra

(F) Self-alignment (medium/slow needs A)

• The goal here is to find regions ion the genome which have multiple approximate copies
• First we need to create local alignments of the genome to itself, then convert it to appropriate format
• Last year done by blastz: https://github.com/fmfi-genomika/genomika-2017/wiki/Self-alignments---self-chain---segmental-duplications
• This yeast I suggest using program last (ubuntu package ast-align, website http://last.cbrc.jp/)
• Here is an example how I have done it in the past, it would be great to rewrite one-liners to some nicer script:

lastdb genome.fa genome.fa 
binqsub -N R-last -l hostname=cpu07 'lastal genome.fa genome.fa -E 1e-20 > self.maf'
maf-convert psl self.maf > tmpC.psl

# filter out trivial self-alignments as well as alignments shorter than 100bp in one of the two sequences or with identity less than 0.9
perl -lane 'die unless @F==21; $s=($F[9] eq $F[13] && $F[10]==$F[12] && $F[11]==0); $s = $s || $F[12]-$F[11]<100 || $F[16]-$F[15]<100 || $F[0]<0.9*($F[0]+$F[1]+$F[2]+$F[3]+$F[5]+$F[7]); print unless $s' tmpC.psl > tmpC100_90.psl
pslToChain tmpC100_90.psl tmpC100_90.chain # kent tools binary, available on genomika
# fix bad coordinates on reverse strand 
perl -lane 'if ($F[0] eq "chain" && $F[9] eq "-") { ($F[10],$F[11]) = ($F[8]-$F[11], $F[8]-$F[10]); print join(" ", @F) } else { print }' tmpC100_90.chain > self100_90.chain

# another chain for alignments with at least 70% identity and length at least 300bo
perl -lane 'die unless @F==21; $s=($F[9] eq $F[13] && $F[10]==$F[12] && $F[11]==0); $s = $s || $F[12]-$F[11]<300 || $F[16]-$F[15]<300 || $F[0]<0.7*($F[0]+$F[1]+$F[2]+$F[3]+$F[5]+$F[7]); print unless $s' tmpC.psl > tmpC300_70.psl
/projects2/dipMag/magCap-2017/assembly/magCapA/seq-tracks/pslToChain tmpC300_70.psl tmpC300_70.chain # kent tools binary copied from genome-dev
# fix bad coordinates on reverse strand 
perl -lane 'if ($F[0] eq "chain" && $F[9] eq "-") { ($F[10],$F[11]) = ($F[8]-$F[11], $F[8]-$F[10]); print join(" ", @F) } else { print }' tmpC300_70.chain > self300_70.chain

Parts of trackDb.ra (replace magCap5 with your genome name):

track selfChain100_90
shortLabel Self aln >90%id
longLabel Self alignments with length >100bp, identity >90%
group varRep
type chain magCapA5

track selfChain300_70
shortLabel Self aln >70%id
longLabel Self alignments with length >300bp, identity >70%
group varRep
type chain magCapA5

(G) Chains between genomes (medium, needs A from both groups)

• TODO: more info

(H) Protein-based chains between genomes (medium, needs A,B from both groups)

• TODO: more info
• https://github.com/fmfi-genomika/genomika-2017/wiki/Chains-from-protein-alignments

(I) Genomes for comparative genomics (fast, only one group)

• Download genomes of additional Malassezia species (other than malGlo and malSym)
• Use list here [3]
• Rename chromosomes similarly as in A, name fasta files in a systematic way (malRes1.fa etc.)
• Store files in a directory at genomika server

(J) Multiple whole-genome alignment (slow, needs A from both groups, I)

• TODO: more info
• https://github.com/fmfi-genomika/genomika-2017/wiki/Alignments

(K) Conservation by phyloP (medium, needs A,I,J)

• TODO: more info
• https://github.com/fmfi-genomika/genomika-2017/wiki/PhyloP-tracks

(L) Conserved elements by phastCons (medium, needs A,I,J)

• TODO: more info
• https://github.com/fmfi-genomika/genomika-2017/wiki/Conservation

(M) Protein domain and other protein annotation from Uniprot (medium, needs A,B)

• TODO: more info
• https://github.com/fmfi-genomika/genomika-2017/wiki/Uniprot-data

(N) Expression data from RNA-seq (medium/slow, needs A)

• TODO: more info
• https://github.com/fmfi-genomika/genomika-2017/wiki/Expression-tracks

(O) Differences between strains (slow, needs A)

• TODO: more info
• https://github.com/fmfi-genomika/genomika-2017/wiki/Strain-comparison