From 697767f1c90470a5b7ed4dd32e3344b52448737c Mon Sep 17 00:00:00 2001 From: Tom Hill Date: Mon, 11 Nov 2024 23:06:23 +1100 Subject: [PATCH] Example formats --- README.md | 10 +++- docs/formats.md | 50 ++++++++++++++++++ docs/manipulating-genome.md | 55 ++++++++++++++++++++ examples/ex1.fa | 56 +++++++++++++++++++++ examples/ex1.sam.gz | Bin 0 -> 114565 bytes examples/ex1_readme.md | 43 ++++++++++++++++ examples/human_g1k_v37.fasta.fai | 84 +++++++++++++++++++++++++++++++ examples/toy.fa | 4 ++ examples/toy.sam | 14 ++++++ 9 files changed, 315 insertions(+), 1 deletion(-) create mode 100644 docs/formats.md create mode 100644 docs/manipulating-genome.md create mode 100644 examples/ex1.fa create mode 100644 examples/ex1.sam.gz create mode 100644 examples/ex1_readme.md create mode 100644 examples/human_g1k_v37.fasta.fai create mode 100644 examples/toy.fa create mode 100644 examples/toy.sam diff --git a/README.md b/README.md index f9a780a..631340c 100644 --- a/README.md +++ b/README.md @@ -3,10 +3,18 @@ DNAvigate is Tom's experimental genome browser, primarily so he can learn about different file formats / methods in bioinformatics-land. +...but its also because no one has named a genome browser DNAvigate yet? And that seems +like the obvious naming choice??? + Please don't expect this to be functional or useful! :) ## Existing Genome Browsers - UCSC genome brower - https://genome.ucsc.edu/ - https://bioconductor.org/packages/release/bioc/vignettes/Gviz/inst/doc/Gviz.html -- IGV - https://igv.org/doc/desktop/ \ No newline at end of file +- IGV - https://igv.org/doc/desktop/ + + +## Stuff I'm Learning about as I build this + +- [file formats](./docs/formats.md) \ No newline at end of file diff --git a/docs/formats.md b/docs/formats.md new file mode 100644 index 0000000..f914f9c --- /dev/null +++ b/docs/formats.md @@ -0,0 +1,50 @@ +# File Formats + +Different file formats in bioinformatics and what they're used for + +## FASTA (.fa) + +Raw nucleotide or protein sequence + +[Example](../examples/mygene.fasta) + +## .fai + +Fasta index file generated by `samtools` for fast access to specific regions + +[Example hg19 index positions for chromosome positions](../examples/human_g1k_v37.fasta.fai) + +## FASTQ (.fq) + +FASTA but with quality scores (I think always a [phred quality score](https://en.wikipedia.org/wiki/Phred_quality_score)?). + +Quality score represents probability that the base was called incorrectly by the sequencer + +```fastq +@SEQ_ID +GATTTGGGGTTCAAAGCAGTATCGATCAAATAGTAAATCCATTTGTTCAACTCACAGTTT ++ +!''*((((***+))%%%++)(%%%%).1***-+*''))**55CCF>>>>>>CCCCCCC65 +``` + +[Example](../examples/HI.4019.002.index_7.ANN0831_R1.fastq) + +## Alignment Formats + +### SAM / BAM (Sequence/Binary Alignment Map) + +Aligns a sequence with a reference sequence. BAM is the binary format. + +[example sam](../examples/toy.sam) + +### CRAM (Compressed Reference-oriented Alignment Map) + +Compressed Sequence Alignment - more compressed than BAM. + +## VCF (Variant Call Format) / BCF (binary format) + +Represents sequence variation (e.g. SNPs, indels, structural variants, alternative alleles) + +## PDB (protein data bank format) + +Representing 3d strucutres of molecules / proteins \ No newline at end of file diff --git a/docs/manipulating-genome.md b/docs/manipulating-genome.md new file mode 100644 index 0000000..bbd405a --- /dev/null +++ b/docs/manipulating-genome.md @@ -0,0 +1,55 @@ +# Steps for manipulating a genome and producing output files + +https://www.biostars.org/p/150010/ (thanks jdimatteo) + +https://web.archive.org/web/20161010092833/http://biobits.org/samtools_primer.html + +https://www.melbournebioinformatics.org.au/tutorials/ + +1. Download hg19 Reference Genome + +```bash +curl -O http://ftp.1000genomes.ebi.ac.uk/vol1/ftp/technical/reference/human_g1k_v37.fasta.fai +curl -O http://ftp.1000genomes.ebi.ac.uk/vol1/ftp/technical/reference/human_g1k_v37.fasta.gz +gunzip human_g1k_v37.fasta.gz +``` + +2. Filter out a single chromosome and index it using `samtools` - suite of tools for interacting w/ the file formats + +`samtools faidx` filters out the single chromosome using the `.fai` index file. + +[Bowtie2](https://github.com/BenLangmead/bowtie2) is a read aligner for aligning a sequence to a reference. +I think `minimap2` is an alternative, modern version. + +```bash +samtools faidx human_g1k_v37.fasta 20 > human_g1k_v37_chr20.fasta +bowtie2-build human_g1k_v37_chr20.fasta homo_chr20 +``` + +3. Simulate a read sample + +[wgsim](https://github.com/lh3/wgsim) simulates sequence reads from a reference genome. +It simulates diloid genomes with SNPs and indel polymorphisms. + +```bash +wgsim -N 1 human_g1k_v37_chr20.fasta single.read1.fq single.read2.fq > wgsim.out +``` + +4. Generate the sam + +```bash +bowtie2 homo_chr20 -1 single.read1.fq -2 single.read2.fq -S single_pair.sam +``` + +5. Generate the bam + +```bash +samtools view -b -S -o single_pair.bam single_pair.sam +``` + +6. Sort and index it + +```bash +samtools sort single_pair.bam single_pair.sorted +samtools index single_pair.sorted.bam +``` \ No newline at end of file diff --git a/examples/ex1.fa b/examples/ex1.fa new file mode 100644 index 0000000..630eb45 --- /dev/null +++ b/examples/ex1.fa @@ -0,0 +1,56 @@ +>seq1 +CACTAGTGGCTCATTGTAAATGTGTGGTTTAACTCGTCCATGGCCCAGCATTAGGGAGCT +GTGGACCCTGCAGCCTGGCTGTGGGGGCCGCAGTGGCTGAGGGGTGCAGAGCCGAGTCAC +GGGGTTGCCAGCACAGGGGCTTAACCTCTGGTGACTGCCAGAGCTGCTGGCAAGCTAGAG +TCCCATTTGGAGCCCCTCTAAGCCGTTCTATTTGTAATGAAAACTATATTTATGCTATTC +AGTTCTAAATATAGAAATTGAAACAGCTGTGTTTAGTGCCTTTGTTCAACCCCCTTGCAA +CAACCTTGAGAACCCCAGGGAATTTGTCAATGTCAGGGAAGGAGCATTTTGTCAGTTACC +AAATGTGTTTATTACCAGAGGGATGGAGGGAAGAGGGACGCTGAAGAACTTTGATGCCCT +CTTCTTCCAAAGATGAAACGCGTAACTGCGCTCTCATTCACTCCAGCTCCCTGTCACCCA +ATGGACCTGTGATATCTGGATTCTGGGAAATTCTTCATCCTGGACCCTGAGAGATTCTGC +AGCCCAGCTCCAGATTGCTTGTGGTCTGACAGGCTGCAACTGTGAGCCATCACAATGAAC +AACAGGAAGAAAAGGTCTTTCAAAAGGTGATGTGTGTTCTCATCAACCTCATACACACAC +ATGGTTTAGGGGTATAATACCTCTACATGGCTGATTATGAAAACAATGTTCCCCAGATAC +CATCCCTGTCTTACTTCCAGCTCCCCAGAGGGAAAGCTTTCAACGCTTCTAGCCATTTCT +TTTGGCATTTGCCTTCAGACCCTACACGAATGCGTCTCTACCACAGGGGGCTGCGCGGTT +TCCCATCATGAAGCACTGAACTTCCACGTCTCATCTAGGGGAACAGGGAGGTGCACTAAT +GCGCTCCACGCCCAAGCCCTTCTCACAGTTTCTGCCCCCAGCATGGTTGTACTGGGCAAT +ACATGAGATTATTAGGAAATGCTTTACTGTCATAACTATGAAGAGACTATTGCCAGATGA +ACCACACATTAATACTATGTTTCTTATCTGCACATTACTACCCTGCAATTAATATAATTG +TGTCCATGTACACACGCTGTCCTATGTACTTATCATGACTCTATCCCAAATTCCCAATTA +CGTCCTATCTTCTTCTTAGGGAAGAACAGCTTAGGTATCAATTTGGTGTTCTGTGTAAAG +TCTCAGGGAGCCGTCCGTGTCCTCCCATCTGGCCTCGTCCACACTGGTTCTCTTGAAAGC +TTGGGCTGTAATGATGCCCCTTGGCCATCACCCAGTCCCTGCCCCATCTCTTGTAATCTC +TCTCCTTTTTGCTGCATCCCTGTCTTCCTCTGTCTTGATTTACTTGTTGTTGGTTTTCTG +TTTCTTTGTTTGATTTGGTGGAAGACATAATCCCACGCTTCCTATGGAAAGGTTGTTGGG +AGATTTTTAATGATTCCTCAATGTTAAAATGTCTATTTTTGTCTTGACACCCAACTAATA +TTTGTCTGAGCAAAACAGTCTAGATGAGAGAGAACTTCCCTGGAGGTCTGATGGCGTTTC +TCCCTCGTCTTCTTA +>seq2 +TTCAAATGAACTTCTGTAATTGAAAAATTCATTTAAGAAATTACAAAATATAGTTGAAAG +CTCTAACAATAGACTAAACCAAGCAGAAGAAAGAGGTTCAGAACTTGAAGACAAGTCTCT +TATGAATTAACCCAGTCAGACAAAAATAAAGAAAAAAATTTTAAAAATGAACAGAGCTTT +CAAGAAGTATGAGATTATGTAAAGTAACTGAACCTATGAGTCACAGGTATTCCTGAGGAA +AAAGAAAAAGTGAGAAGTTTGGAAAAACTATTTGAGGAAGTAATTGGGGAAAACCTCTTT +AGTCTTGCTAGAGATTTAGACATCTAAATGAAAGAGGCTCAAAGAATGCCAGGAAGATAC +ATTGCAAGACAGACTTCATCAAGATATGTAGTCATCAGACTATCTAAAGTCAACATGAAG +GAAAAAAATTCTAAAATCAGCAAGAGAAAAGCATACAGTCATCTATAAAGGAAATCCCAT +CAGAATAACAATGGGCTTCTCAGCAGAAACCTTACAAGCCAGAAGAGATTGGATCTAATT +TTTGGACTTCTTAAAGAAAAAAAAACCTGTCAAACACGAATGTTATGCCCTGCTAAACTA +AGCATCATAAATGAAGGGGAAATAAAGTCAAGTCTTTCCTGACAAGCAAATGCTAAGATA +ATTCATCATCACTAAACCAGTCCTATAAGAAATGCTCAAAAGAATTGTAAAAGTCAAAAT +TAAAGTTCAATACTCACCATCATAAATACACACAAAAGTACAAAACTCACAGGTTTTATA +AAACAATTGAGACTACAGAGCAACTAGGTAAAAAATTAACATTACAACAGGAACAAAACC +TCATATATCAATATTAACTTTGAATAAAAAGGGATTAAATTCCCCCACTTAAGAGATATA +GATTGGCAGAACAGATTTAAAAACATGAACTAACTATATGCTGTTTACAAGAAACTCATT 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zxp7_Nx(900?gBr<_Q-GjleXwK!XlPykD@+K?qBg>Fo08Kskc5UG!|*~+#-`V7Z(mAZ21`4s*kn$)_(&B8 z62*2LWk(Y-Imn&$PM7kZubHav<7V_p2O;gsK{`1&b-zEIt@z4)<{3>iZbsi8BBRBw z)TIvk_pUQIpQ%Z(d6w+S{E>Od-`<{{qU&`Qb81((uF@6FFVeX_y2GS|zBc3*<>&6f zNhGY3Pw#)fbiaO|qLe!h_P)jxJ3gLKi6J ex1.vcf + +Generate a compressed VCF file of variants: + samtools mpileup -g -f ex1.fa ex1.bam > ex1.bcf \ No newline at end of file diff --git a/examples/human_g1k_v37.fasta.fai b/examples/human_g1k_v37.fasta.fai new file mode 100644 index 0000000..3ea5cac --- /dev/null +++ b/examples/human_g1k_v37.fasta.fai @@ -0,0 +1,84 @@ +1 249250621 52 60 61 +2 243199373 253404903 60 61 +3 198022430 500657651 60 61 +4 191154276 701980507 60 61 +5 180915260 896320740 60 61 +6 171115067 1080251307 60 61 +7 159138663 1254218344 60 61 +8 146364022 1416009371 60 61 +9 141213431 1564812846 60 61 +10 135534747 1708379889 60 61 +11 135006516 1846173603 60 61 +12 133851895 1983430282 60 61 +13 115169878 2119513096 60 61 +14 107349540 2236602526 60 61 +15 102531392 2345741279 60 61 +16 90354753 2449981581 60 61 +17 81195210 2541842300 60 61 +18 78077248 2624390817 60 61 +19 59128983 2703769406 60 61 +20 63025520 2763883926 60 61 +21 48129895 2827959925 60 61 +22 51304566 2876892038 60 61 +X 155270560 2929051733 60 61 +Y 59373566 3086910193 60 61 +MT 16569 3147273397 70 71 +GL000207.1 4262 3147290265 60 61 +GL000226.1 15008 3147294661 60 61 +GL000229.1 19913 3147309982 60 61 +GL000231.1 27386 3147330289 60 61 +GL000210.1 27682 3147358194 60 61 +GL000239.1 33824 3147386400 60 61 +GL000235.1 34474 3147420850 60 61 +GL000201.1 36148 3147455961 60 61 +GL000247.1 36422 3147492774 60 61 +GL000245.1 36651 3147529866 60 61 +GL000197.1 37175 3147567190 60 61 +GL000203.1 37498 3147605047 60 61 +GL000246.1 38154 3147643232 60 61 +GL000249.1 38502 3147682084 60 61 +GL000196.1 38914 3147721290 60 61 +GL000248.1 39786 3147760915 60 61 +GL000244.1 39929 3147801427 60 61 +GL000238.1 39939 3147842084 60 61 +GL000202.1 40103 3147882751 60 61 +GL000234.1 40531 3147923585 60 61 +GL000232.1 40652 3147964854 60 61 +GL000206.1 41001 3148006246 60 61 +GL000240.1 41933 3148047993 60 61 +GL000236.1 41934 3148090687 60 61 +GL000241.1 42152 3148133382 60 61 +GL000243.1 43341 3148176299 60 61 +GL000242.1 43523 3148220425 60 61 +GL000230.1 43691 3148264736 60 61 +GL000237.1 45867 3148309218 60 61 +GL000233.1 45941 3148355912 60 61 +GL000204.1 81310 3148402681 60 61 +GL000198.1 90085 3148485409 60 61 +GL000208.1 92689 3148577058 60 61 +GL000191.1 106433 3148671355 60 61 +GL000227.1 128374 3148779625 60 61 +GL000228.1 129120 3148910202 60 61 +GL000214.1 137718 3149041537 60 61 +GL000221.1 155397 3149181614 60 61 +GL000209.1 159169 3149339664 60 61 +GL000218.1 161147 3149501549 60 61 +GL000220.1 161802 3149665445 60 61 +GL000213.1 164239 3149830007 60 61 +GL000211.1 166566 3149997047 60 61 +GL000199.1 169874 3150166453 60 61 +GL000217.1 172149 3150339222 60 61 +GL000216.1 172294 3150514304 60 61 +GL000215.1 172545 3150689533 60 61 +GL000205.1 174588 3150865017 60 61 +GL000219.1 179198 3151042578 60 61 +GL000224.1 179693 3151224826 60 61 +GL000223.1 180455 3151407577 60 61 +GL000195.1 182896 3151591103 60 61 +GL000212.1 186858 3151777111 60 61 +GL000222.1 186861 3151967147 60 61 +GL000200.1 187035 3152157186 60 61 +GL000193.1 189789 3152347402 60 61 +GL000194.1 191469 3152540418 60 61 +GL000225.1 211173 3152735142 60 61 +GL000192.1 547496 3152949898 60 61 diff --git a/examples/toy.fa b/examples/toy.fa new file mode 100644 index 0000000..ec99cf7 --- /dev/null +++ b/examples/toy.fa @@ -0,0 +1,4 @@ +>ref +AGCATGTTAGATAAGATAGCTGTGCTAGTAGGCAGTCAGCGCCAT +>ref2 +aggttttataaaacaattaagtctacagagcaactacgcg \ No newline at end of file diff --git a/examples/toy.sam b/examples/toy.sam new file mode 100644 index 0000000..0112f61 --- /dev/null +++ b/examples/toy.sam @@ -0,0 +1,14 @@ +@SQ SN:ref LN:45 +@SQ SN:ref2 LN:40 +r001 163 ref 7 30 8M4I4M1D3M = 37 39 TTAGATAAAGAGGATACTG * XX:B:S,12561,2,20,112 +r002 0 ref 9 30 1S2I6M1P1I1P1I4M2I * 0 0 AAAAGATAAGGGATAAA * +r003 0 ref 9 30 5H6M * 0 0 AGCTAA * +r004 0 ref 16 30 6M14N1I5M * 0 0 ATAGCTCTCAGC * +r003 16 ref 29 30 6H5M * 0 0 TAGGC * +r001 83 ref 37 30 9M = 7 -39 CAGCGCCAT * +x1 0 ref2 1 30 20M * 0 0 aggttttataaaacaaataa ???????????????????? +x2 0 ref2 2 30 21M * 0 0 ggttttataaaacaaataatt ????????????????????? +x3 0 ref2 6 30 9M4I13M * 0 0 ttataaaacAAATaattaagtctaca ?????????????????????????? +x4 0 ref2 10 30 25M * 0 0 CaaaTaattaagtctacagagcaac ????????????????????????? +x5 0 ref2 12 30 24M * 0 0 aaTaattaagtctacagagcaact ???????????????????????? +x6 0 ref2 14 30 23M * 0 0 Taattaagtctacagagcaacta ??????????????????????? \ No newline at end of file