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Genomics_Research

This readme was written as instructions for another researcher to pick up on the work I left off.

Step 1: Referencing hg19

hgrefFile = "refGene_hg19_sorted.txt" #prebuilt file  
combine_patient_data_hgRef.build(hgrefFile)

first 5 lines of hg19:

585	NR_024540	chr1	-	14361	29370	29370	29370	11	14361,14969,15795,16606,16857,17232,17605,17914,18267,24737,29320,	14829,15038,15947,16765,17055,17368,17742,18061,18366,24891,29370,	0	WASH7P	unk	unk	-1,-1,-1,-1,-1,-1,-1,-1,-1,-1,-1,  
585	NR_026818	chr1	-	34610	36081	36081	36081	3	34610,35276,35720,	35174,35481,36081,	0	FAM138A	unk	unk	-1,-1,-1,  
585	NR_026820	chr1	-	34610	36081	36081	36081	3	34610,35276,35720,	35174,35481,36081,	0	FAM138F	unk	unk	-1,-1,-1,  
585	NM_001005484	chr1	+	69090	70008	69090	70008	1	69090,	70008,	0	OR4F5	cmpl	cmpl	0,  
587	NR_028322	chr1	+	323891	328581	328581	328581	3	323891,324287,324438,	324060,324345,328581,	0	LOC100132287	unk	unk	-1,-1,-1,

Description of columns can be read up on here
https://genome.ucsc.edu/cgi-bin/hgTables?db=hg19&hgta_group=genes&hgta_track=refGene&hgta_table=refGene&hgta_doSchema=describe+table+schema
We only need the geneId, chromosome number, strand, transcription start and stop points
columns 1,2,3,4,5 respectively

combine_patient_data_hgRef.build() uses the prebuilt patient files (inside the directory "patient_data"), labeled as

ff870342-f0d6-4450-8f9c-344c046a0baf.pcawg_consensus_1.6.161116.somatic.sv.bedpe
ffa976f0-aa60-4867-842e-361afa7d68ac.pcawg_consensus_1.6.161116.somatic.sv.bedpe
ffad9288-c622-11e3-bf01-24c6515278c0.pcawg_consensus_1.6.161116.somatic.sv.bedpe
.
.
.

(those file prefixes (up to .pcawg) will be used later)
and we create a new directory, called "Referenced_patient_data" inside your current working directory. After this process runs (for the size of this data, it takes approximately 10-12 minutes), the "Referenced_patient_data" directory will contain individual files with the same file prefix as the corresponding patient data, but with only the valid gene fusions listed. For example:

#first 5 lines (of ~50 lines) of 0bd3a230-531e-44aa-9999-b8ed8da0176b.pcawg_consensus_1.6.161116.somatic.sv.bedpe
chrom1	start1	end1	chrom2	start2	end2	sv_id	pe_support	strand1	strand2	svclass	svmethod
11	82798867	82798868	11	82936093	82936094	SVMERGE10	37	+	-	DEL	SNOWMAN_BRASS_dRANGER_DELLY
12	115046363	115046364	4	56334945	56334946	SVMERGE34	35	-	+	TRA	SNOWMAN_BRASS_dRANGER_DELLY
12	115046403	115046404	4	72591306	72591307	SVMERGE42	32	+	-	TRA	SNOWMAN_BRASS_dRANGER_DELLY
12	118147063	118147064	13	54442327	54442328	SVMERGE38	12	-	-	TRA	SNOWMAN_BRASS_DELLY

produces

geneA	geneA_orientation	geneB	geneB_orientation	chr1	position	chr1_orientation	chr2	position	chr2_orientation
./patient_data/0bd3a230-531e-44aa-9999-b8ed8da0176b.pcawg_consensus_1.6.161116.somatic.sv.bedpe									
KSR2	-	PCDH9	-	chr12	118147303	+	chr13	67542094	-
CLDN14	-	DYRK1A	+	chr21	37873660	-	chr21	38850309	-
DYRK1A	+	TMPRSS2	-	chr21	38846232	-	chr21	42868379	-
DYRK1A	+	TMPRSS2	-	chr21	38846936	+	chr21	42869068	+
DYRK1A	+	ERG	-	chr21	38850232	+	chr21	39893645	+
AFF1	+	INTS12	-	chr4	88026639	-	chr4	106603789	-
INTS12	-	KIAA1429	-	chr4	106603808	+	chr8	95541749	-

That's the entire tsv file. First line column headers, second line is the original source file, followed by the data
So looking at this example, this patient has 7 gene fusions.
The first one contains genes KSR2 and PCDH9 with the breakpoint on KSR2 located at 118147303 and the breakpoint on PDCH9 located on 67542094. These locations will be used later.

Step 2 Creating a lot of files

This step uses all of the following prexisting files. (first 5 lines included)

gencode.v19.annotation.gtf_withproteinids (approx. 1.1 GB)

#first 5 lines
chr1	HAVANA	gene	11869	14412	.	+	.	gene_id "ENSG00000223972.4"; transcript_id "ENSG00000223972.4"; gene_type "pseudogene"; gene_status "KNOWN"; gene_name "DDX11L1"; transcript_type "pseudogene"; transcript_status "KNOWN"; transcript_name "DDX11L1"; level 2; havana_gene "OTTHUMG00000000961.2";
chr1	HAVANA	transcript	11869	14409	.	+	.	gene_id "ENSG00000223972.4"; transcript_id "ENST00000456328.2"; gene_type "pseudogene"; gene_status "KNOWN"; gene_name "DDX11L1"; transcript_type "processed_transcript"; transcript_status "KNOWN"; transcript_name "DDX11L1-002"; level 2; tag "basic"; havana_gene "OTTHUMG00000000961.2"; havana_transcript "OTTHUMT00000362751.1";
chr1	HAVANA	exon	11869	12227	.	+	.	gene_id "ENSG00000223972.4"; transcript_id "ENST00000456328.2"; gene_type "pseudogene"; gene_status "KNOWN"; gene_name "DDX11L1"; transcript_type "processed_transcript"; transcript_status "KNOWN"; transcript_name "DDX11L1-002"; exon_number 1; exon_id "ENSE00002234944.1"; level 2; tag "basic"; havana_gene "OTTHUMG00000000961.2"; havana_transcript "OTTHUMT00000362751.1";
chr1	HAVANA	exon	12613	12721	.	+	.	gene_id "ENSG00000223972.4"; transcript_id "ENST00000456328.2"; gene_type "pseudogene"; gene_status "KNOWN"; gene_name "DDX11L1"; transcript_type "processed_transcript"; transcript_status "KNOWN"; transcript_name "DDX11L1-002"; exon_number 2; exon_id "ENSE00003582793.1"; level 2; tag "basic"; havana_gene "OTTHUMG00000000961.2"; havana_transcript "OTTHUMT00000362751.1";
chr1	HAVANA	exon	13221	14409	.	+	.	gene_id "ENSG00000223972.4"; transcript_id "ENST00000456328.2"; gene_type "pseudogene"; gene_status "KNOWN"; gene_name "DDX11L1"; transcript_type "processed_transcript"; transcript_status "KNOWN"; transcript_name "DDX11L1-002"; exon_number 3; exon_id "ENSE00002312635.1"; level 2; tag "basic"; havana_gene "OTTHUMG00000000961.2"; havana_transcript "OTTHUMT00000362751.1";

Getting a better understanding of this file is important

You'll need to talk to Dr. Yang about this file.   The issue is this:
You see how the 3rd entry in each line is "gene", "transcript", "exon", "exon", "exon" in that order?
So you'll see later that we're looking at the particular RNA-seq value for each exon. However, in the later file that has that data, the exons are labeled by exon number, which, if you ctrl-f "exon_number", you'll see is contained in each line that starts with "exon". However(!), it doesn't occur in this subset, but occassionally you'll get exons on the same gene that have the same number. There's an attribute called exon_id, unfortunately, the RNA-seq value file doesn't list by id. The reason this is a problem is because, while the RNA-seq file will just have one value for one exon number (per patient), the location of that exon (cited in this file) varies for different exons, which will affect which side of the gene breakpoint they are, and therefore affect the results of your data. That's the best I can explain it over text, Dr. Yang can help you further with this.

counts_all_2015_combined_exon_dex.STAR.whitelist.tsv (approx. 2.7 GB)

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Each column in this file corresponds to one patient file
each row in the first column is a specific exon on that geneId (labeled ENSG***)
and then every value in the cells below a particular patient is the RNA-seq value on that exon on that gene for that patient. This is the focus of the analysis, but there's a lot more file leg work that needs to be done.

Referenced_patient_data

This is just the previously build directory of patient gene-fusion files

pcawg_summary.tsv

donor_unique_id	wgs_exclusion_white_gray	dcc_project_code	submitter_donor_id	icgc_donor_id	mar2016_donor	santa_cruz_pilot	validation_by_deep_seq	TiN	normal_wgs_submitter_specimen_id	normal_wgs_icgc_specimen_id	normal_wgs_submitter_sample_id	normal_wgs_icgc_sample_id	normal_wgs_aliquot_id	normal_wgs_bwa_alignment_gnos_repo	normal_wgs_bwa_alignment_gnos_id	is_mar2016_normal_wgs_bwa_alignment	normal_wgs_bwa_alignment_bam_file_name	normal_wgs_minibam_gnos_repo	normal_wgs_minibam_gnos_id	is_mar2016_normal_wgs_minibam	normal_wgs_minibam_bam_file_name	tumor_wgs_specimen_count	tumor_wgs_submitter_specimen_id	tumor_wgs_icgc_specimen_id	tumor_wgs_submitter_sample_id	tumor_wgs_icgc_sample_id	tumor_wgs_aliquot_idtumor_wgs_oxog_score	tumor_wgs_ContEST	tumor_wgs_Stars	tumor_wgs_bwa_alignment_gnos_repo	tumor_wgs_bwa_alignment_gnos_id	is_mar2016_tumor_wgs_bwa_alignment	tumor_wgs_bwa_alignment_bam_file_name	tumor_wgs_minibam_gnos_repo	tumor_wgs_minibam_gnos_id	is_mar2016_tumor_wgs_minibam	tumor_wgs_minibam_bam_file_name	sanger_variant_calling_repo	sanger_variant_calling_gnos_id	sanger_variant_calling_file_name_prefix	is_mar2016_sanger_variant_calling	sanger_variant_calling_deprecated_gnos_id	dkfz_embl_variant_calling_repo	dkfz_embl_variant_calling_gnos_id	dkfz_embl_variant_calling_file_name_prefix	is_mar2016_dkfz_embl_variant_calling	dkfz_embl_variant_calling_deprecated_gnos_id	broad_variant_calling_repo	broad_variant_calling_gnos_id	broad_variant_calling_file_name_prefix	is_mar2016_broad_variant_calling	broad_variant_calling_deprecated_gnos_id	muse_variant_calling_repo	muse_variant_calling_gnos_id	muse_variant_calling_file_name_prefix	is_mar2016_muse_variant_calling	broad_tar_variant_calling_repo	broad_tar_variant_calling_gnos_id	broad_tar_variant_calling_file_name_prefix	is_mar2016_broad_tar_variant_calling	normal_wgs_has_matched_rna_seq	tumor_wgs_has_matched_rna_seq	normal_rna_seq_submitter_specimen_id	normal_rna_seq_icgc_specimen_id	normal_rna_seq_submitter_sample_id	normal_rna_seq_icgc_sample_id	normal_rna_seq_aliquot_id	normal_rna_seq_star_alignment_gnos_repo	normal_rna_seq_star_alignment_gnos_id	is_mar2016_normal_rna_seq_star_alignment	normal_rna_seq_star_alignment_bam_file_name	normal_rna_seq_tophat_alignment_gnos_repo	normal_rna_seq_tophat_alignment_gnos_id	is_mar2016_normal_rna_seq_tophat_alignment	normal_rna_seq_tophat_alignment_bam_file_name	tumor_rna_seq_specimen_count	tumor_rna_seq_submitter_specimen_id	tumor_rna_seq_icgc_specimen_id	tumor_rna_seq_submitter_sample_id	tumor_rna_seq_icgc_sample_idtumor_rna_seq_aliquot_id	tumor_rna_seq_star_alignment_gnos_repo	tumor_rna_seq_star_alignment_gnos_idis_mar2016_tumor_rna_seq_star_alignment	tumor_rna_seq_star_alignment_bam_file_name	tumor_rna_seq_tophat_alignment_gnos_repo	tumor_rna_seq_tophat_alignment_gnos_id	is_mar2016_tumor_rna_seq_tophat_alignment	tumor_rna_seq_tophat_alignment_bam_file_name
BLCA-US::096b4f32-10c1-4737-a0dd-cae04c54ee33	Whitelist	BLCA-US	096b4f32-10c1-4737-a0dd-cae04c54ee33	DO804	True	True	True	1.000000e-02	27461a27-26eb-4c2c-9c54-e16fbd32c615	SP1682	e0fccaf5-925a-41f9-b87c-cd5ee4aecb59	SA5237	e0fccaf5-925a-41f9-b87c-cd5ee4aecb59	https://gtrepo-osdc-tcga.annailabs.com/	f22a72c5-73c8-478d-b03e-04599b9d5321	True	PCAWG.67455c36-aa47-4cc4-8b6d-9a9012b616ed.bam	https://gtrepo-osdc-tcga.annailabs.com/	7e442040-f6ea-11e5-bd75-997f5888a0e8	False	PCAWG.67455c36-aa47-4cc4-8b6d-9a9012b616ed_minibam.bam	1	52f538ef-b05d-4c76-9976-ce6d49158016	SP1677	301d6ce3-4099-4c1d-8e50-c04b7ce91450	SA5195	301d6ce3-4099-4c1d-8e50-c04b7ce91450	87.52	0.1	5.0	https://gtrepo-osdc-tcga.annailabs.com/	a6e8dd23-c8a5-445a-ae4b-b9f92ed6a73e	True	PCAWG.a833602e-7ee7-442b-9bf9-f6489cd64ba2.bam	https://gtrepo-osdc-tcga.annailabs.com/	7e442040-f6ea-11e5-bd75-997f5888a0e8	False	PCAWG.a833602e-7ee7-442b-9bf9-f6489cd64ba2_minibam.bam	https://gtrepo-osdc-tcga.annailabs.com/	8935fe36-ceec-11e5-bedc-767d4d100685301d6ce3-4099-4c1d-8e50-c04b7ce91450	True	5230346f-987e-44c3-b9fe-dddb5829e247	https://gtrepo-osdc-tcga.annailabs.com/	4269588d-5012-4eee-ab9a-9c48897dae69	301d6ce3-4099-4c1d-8e50-c04b7ce91450	True		https://gtrepo-osdc-tcga.annailabs.com/	ac34e35d-21b1-4b11-b42d-155cba1a547f301d6ce3-4099-4c1d-8e50-c04b7ce91450	True	d12904ff-4ce6-4d7b-a0d7-69e18a3822cc	https://gtrepo-osdc-tcga.annailabs.com/	48077833-99ed-49c6-88a1-80cd2e50755a	301d6ce3-4099-4c1d-8e50-c04b7ce91450	True	https://gtrepo-osdc-tcga.annailabs.com/	38ce6165-c324-4642-89cf-984d738ee08a	301d6ce3-4099-4c1d-8e50-c04b7ce91450	True	False	True							152f538ef-b05d-4c76-9976-ce6d49158016	SP1677	22e154de-0e3b-443b-8420-48d68d6c1ce4	SA5213	22e154de-0e3b-443b-8420-48d68d6c1ce4	https://gtrepo-osdc-tcga.annailabs.com/	a63c9c0a-c8aa-44dc-8915-08f55109a5c9	True	PCAWG.f4288632-a94f-4777-a5d6-1a76b12abcc5.STAR.v1.bam	https://gtrepo-osdc-tcga.annailabs.com/	81438f7c-ee87-44a7-acd1-562ec8f386f0	True	PCAWG.f4288632-a94f-4777-a5d6-1a76b12abcc5.TopHat2.v1.bam
BLCA-US::178b28cd-99c3-48dc-8d09-1ef71b4cee80	Whitelist	BLCA-US	178b28cd-99c3-48dc-8d09-1ef71b4cee80	DO555	True	True	True	0	cd3cfb26-e66f-408e-81f6-3b61c247c976	SP1135	c1da8eed-4919-4ba5-a735-3fba476c18a7	SA1598	c1da8eed-4919-4ba5-a735-3fba476c18a7	https://gtrepo-osdc-tcga.annailabs.com/	964bac8f-43cc-4a47-b233-53798d287029	True	PCAWG.13985b6b-d7b6-4efd-9c53-288f4d269a0c.bam	https://gtrepo-osdc-tcga.annailabs.com/	0b6a5d16-e617-11e5-ad33-d98536d081eeFalse	PCAWG.13985b6b-d7b6-4efd-9c53-288f4d269a0c_minibam.bam	1	59d6683f-5eb7-493d-8e8e-78b88be2cd70	SP1132	4838b5a9-968c-4178-bffb-3fafe1f6dc09	SA1556	4838b5a9-968c-4178-bffb-3fafe1f6dc0954.55	0.1	5.0	https://gtrepo-osdc-tcga.annailabs.com/	437e11a0-4137-4614-9f64-c5e798c8bb33True	PCAWG.af830047-822a-4dcc-88bc-68f7440a208d.bam	https://gtrepo-osdc-tcga.annailabs.com/	0b6a5d16-e617-11e5-ad33-d98536d081ee	False	PCAWG.af830047-822a-4dcc-88bc-68f7440a208d_minibam.bam	https://gtrepo-osdc-tcga.annailabs.com/	714e7b6a-d34a-11e5-90f0-f8135c6878de	4838b5a9-968c-4178-bffb-3fafe1f6dc09	True	c0387aa4-9937-45f7-bf3a-e2da65457738	https://gtrepo-osdc-tcga.annailabs.com/	875fdb54-76ae-11e5-abee-bff16f24f4a1	4838b5a9-968c-4178-bffb-3fafe1f6dc09	True		https://gtrepo-osdc-tcga.annailabs.com/	d989e42d-354c-4652-9921-27f79ba31c2e	4838b5a9-968c-4178-bffb-3fafe1f6dc09	True	f3c6d06b-09ff-4e0d-821d-80bd2b04106f	https://gtrepo-osdc-tcga.annailabs.com/	d9ec79b7-0ccb-4312-b84d-b90200eb77bf	4838b5a9-968c-4178-bffb-3fafe1f6dc09	True	https://gtrepo-osdc-tcga.annailabs.com/	9f6910ec-80dc-4270-b415-26c00f5ad281	4838b5a9-968c-4178-bffb-3fafe1f6dc09	True	False	True									159d6683f-5eb7-493d-8e8e-78b88be2cd70	SP1132	f154cc00-e118-4d4c-b9d2-2f1f71e0a964	SA1580	f154cc00-e118-4d4c-b9d2-2f1f71e0a964	https://gtrepo-osdc-tcga.annailabs.com/	2c4bd7a6-721a-4cd1-aa40-90cecb01c32c	True	PCAWG.5c2ab9dd-c1ed-4d57-b68d-56a7b52b80e0.STAR.v1.bam	https://gtrepo-osdc-tcga.annailabs.com/	4a81f567-fb9b-4561-b2b7-c099c206a2ce	True	PCAWG.5c2ab9dd-c1ed-4d57-b68d-56a7b52b80e0.TopHat2.v1.bam
BLCA-US::1e308b12-0590-4dae-94d0-a539fcf25df7	Whitelist	BLCA-US	1e308b12-0590-4dae-94d0-a539fcf25df7	DO479	True	True	True	0	59b99c9a-d1c0-4cc0-953b-4358bcf9f392	SP968	d6e91f4c-38c0-4393-9cb7-be076663dff3	SA3784	d6e91f4c-38c0-4393-9cb7-be076663dff3	https://gtrepo-osdc-tcga.annailabs.com/	fa36e89d-55b8-4c72-9392-80113efde076	True	PCAWG.c878fe80-0a2e-43d2-ab62-1214fc34c282.bam	https://gtrepo-osdc-tcga.annailabs.com/	58624910-e5a1-11e5-8a85-b49cebf72b8fFalse	PCAWG.c878fe80-0a2e-43d2-ab62-1214fc34c282_minibam.bam	1	e9889071-4c6e-4761-9d65-06c6b5989fb7	SP967	c66c92d5-df65-46e6-861d-d8a98808e6a3	SA3742	c66c92d5-df65-46e6-861d-d8a98808e6a386.05	0.1	5.0	https://gtrepo-osdc-tcga.annailabs.com/	416911eb-e10f-4edd-8f07-5e87b0228a11True	PCAWG.066b5025-1a70-4db4-bd98-964b8c9939c3.bam	https://gtrepo-osdc-tcga.annailabs.com/	58624910-e5a1-11e5-8a85-b49cebf72b8f	False	PCAWG.066b5025-1a70-4db4-bd98-964b8c9939c3_minibam.bam	https://gtrepo-osdc-tcga.annailabs.com/	67959d40-d34c-11e5-bc3a-f8135c6878de	c66c92d5-df65-46e6-861d-d8a98808e6a3	True	838819c6-29d1-4d45-b534-d198dfffdaa9	https://gtrepo-osdc-tcga.annailabs.com/	a44f0d88-76af-11e5-b61f-84cb5485c5b1	c66c92d5-df65-46e6-861d-d8a98808e6a3	True		https://gtrepo-osdc-tcga.annailabs.com/	5c58c28e-b8b8-4137-bf8d-526fffab8338	c66c92d5-df65-46e6-861d-d8a98808e6a3	True	7e45c6f6-0b5c-474f-b251-2bf58be35290	https://gtrepo-osdc-tcga.annailabs.com/	854c515b-bade-4f10-8f24-a46b0abd1507	c66c92d5-df65-46e6-861d-d8a98808e6a3	True	https://gtrepo-osdc-tcga.annailabs.com/	595384e4-c0e1-47d2-a1e3-c1aa7f9eeea6	c66c92d5-df65-46e6-861d-d8a98808e6a3	True	False	True									1e9889071-4c6e-4761-9d65-06c6b5989fb7	SP967	429f0e9d-c160-4b36-a31b-dab1097c6129	SA3760	429f0e9d-c160-4b36-a31b-dab1097c6129	https://gtrepo-osdc-tcga.annailabs.com/	5e5f155b-0c8c-4917-b47c-606fda32b0e8	True	PCAWG.ba7d499b-049b-4ba2-be02-e513f81d453e.STAR.v1.bam	https://gtrepo-osdc-tcga.annailabs.com/	4c45ff2a-309f-4167-936e-e5b894ce6b79	True	PCAWG.ba7d499b-049b-4ba2-be02-e513f81d453e.TopHat2.v1.bam
BLCA-US::24f21425-b001-4986-aedf-5b4dd851c6ad	Whitelist	BLCA-US	24f21425-b001-4986-aedf-5b4dd851c6ad	DO689	True	True	False	1.000000e-02	1c7630ed-6b1c-4569-a15c-e296772868de	SP1425	f88b2f4c-15f1-4808-bcec-89abc7466de6	SA3769	f88b2f4c-15f1-4808-bcec-89abc7466de6	https://gtrepo-osdc-tcga.annailabs.com/	b2a9fda3-15e8-4a8b-8a8a-195d72573423	True	PCAWG.c3ffa75b-de27-4cc6-b713-b32c51f5bee5.bam	https://gtrepo-osdc-tcga.annailabs.com/	3456a156-e8ee-11e5-bd56-a21f777315da	False	PCAWG.c3ffa75b-de27-4cc6-b713-b32c51f5bee5_minibam.bam	1	967443e3-35c3-4f41-9bb5-dc899d290806	SP1419	973d0577-8ca4-44a1-817f-1d3c1bada151	SA3721	973d0577-8ca4-44a1-817f-1d3c1bada151	87.09	0.1	4.5	https://gtrepo-osdc-tcga.annailabs.com/	d2e25eb8-8899-4ba5-a2f1-0a69ba97cef2	True	PCAWG.bd9f7681-6ed4-4f60-8db9-eb7ab0d4a7e7.bam	https://gtrepo-osdc-tcga.annailabs.com/	3456a156-e8ee-11e5-bd56-a21f777315da	False	PCAWG.bd9f7681-6ed4-4f60-8db9-eb7ab0d4a7e7_minibam.bam	https://gtrepo-osdc-tcga.annailabs.com/	bd31974a-cfdc-11e5-b3ed-eaf5899203f5973d0577-8ca4-44a1-817f-1d3c1bada151	True	d1fc904c-eb10-4d61-adea-58fb70412b7a	https://gtrepo-osdc-tcga.annailabs.com/	4508880d-8850-471e-9170-dddff85fd241	973d0577-8ca4-44a1-817f-1d3c1bada151	True		https://gtrepo-osdc-tcga.annailabs.com/	927237c0-427d-4182-9023-fa42b94c6d3b973d0577-8ca4-44a1-817f-1d3c1bada151	True	6f399d0f-e0b4-4ae2-9d2a-e7c141825708	https://gtrepo-osdc-tcga.annailabs.com/	239d1ef8-53b5-4571-a663-cfc651ec2e8a	973d0577-8ca4-44a1-817f-1d3c1bada151	True	https://gtrepo-osdc-tcga.annailabs.com/	4c1298ba-2a99-4e2c-859d-b0c7dec2fce0	973d0577-8ca4-44a1-817f-1d3c1bada151	True	False	True							1967443e3-35c3-4f41-9bb5-dc899d290806	SP1419	80cdd385-2a6a-4d59-81c7-f36786637323	SA3745	80cdd385-2a6a-4d59-81c7-f36786637323	https://gtrepo-osdc-tcga.annailabs.com/	527cc5b2-8716-44ae-9d63-8f16c4857184	True	PCAWG.65e2deab-5ba3-4d8c-83a9-69b29e0e8e40.STAR.v1.bam	https://gtrepo-osdc-tcga.annailabs.com/	90f6db49-d0e0-4b2b-b859-01fe4b90faaa	True	PCAWG.65e2deab-5ba3-4d8c-83a9-69b29e0e8e40.TopHat2.v1.bam

This is the file that references those long complicated fileheaders from earlier to a specific patient id.
We need this because we also have...

./exonRefs/pcawg.rnaseq.extended.metadata.aliquot_id.V4.tsv

aliquot_id	study	project_code	icgc_donor_id	icgc_specimen_id	icgc_sample_id	submitted_donor_id	submitted_specimen_id	submitted_sample_id	specimen_type	donor_id	specimen_id	sample_id	date	file_name	filesize	md5_checksum	analysis_id	uri	center_name	platform	platform_model	lib_id	rg_label	files	files_blacklist	pilot_63	includes_spike_ins	spike_ins_fasta	spike_ins_concentration	library_type	library_strategy	accession	state	is_tumour	file_type	unique_donor_id	wgs_white_black_graywgs_exclusion_white_gray	donor_unique_id	tcga_donor_uuid	donor_sex	donor_vital_status	donor_diagnosis_icd10	first_therapy_type	first_therapy_response	donor_age_at_diagnosis	donor_survival_time	donor_interval_of_last_followup	tobacco_smoking_history_indicator	tobacco_smoking_intensity	alcohol_history	alcohol_history_intensity	donor_wgs_included_excluded	tcga_specimen_uuid	tcga_sample_uuid	organ_system	histology_abbreviation	histology_tier1	histology_tier2	histology_tier3	histology_tier4	tumour_histological_code	tumour_histological_type	tumour_stage	tumour_grade	percentage_cellularity	level_of_cellularity	tcga_expert_re.review	tumour_histological_comment	specimen_donor_treatment_type	rna_seq_specimen_id	Color (RGB code)	rna_seq_aliquot_id	wgs_aliquot_id	GTex.tissue	tumor.normal	matched_wgs_aliquot_id
b337121c-9821-4644-820e-b8c477f6c38a	Brain Glioblastoma Multiforme - TCGA, US	GBM-US	DO10908	SP23639	SA136044	737b35e1-d668-4fce-9b6e-76946c7952b6	96881d37-7a3e-4910-b897-19453118fe7c	b337121c-9821-4644-820e-b8c477f6c38a	Primary tumour - solid tissue	737b35e1-d668-4fce-9b6e-76946c7952b6	96881d37-7a3e-4910-b897-19453118fe7c	TCGA-27-2523-01A-01R-1850-01	2013-12-17T00:55:04Z	G17483.TCGA-27-2523-01A-01R-1850-01.2.tar.gz	6850833438	d443ea4bbd917b43bd3bc20882f07eb2	52918733-0b2d-462b-8784-70bd148d9ffe	https://cghub.ucsc.edu/cghub/data/analysis/download/52918733-0b2d-462b-8784-70bd148d9ffe	BI	ILLUMINA	Illumina HiSeq 2000	7290.WR29324.Solexa-91309.C0HTRACXX120406.P	C0HTR.3	G17483.TCGA-27-2523-01A-01R-1850-01.2.bam	NA	FALSENA	NA		RNA-Seq			yes	fastq	GBM-US::737b35e1-d668-4fce-9b6e-76946c7952b6	Whitelist	Whitelist	GBM-US::737b35e1-d668-4fce-9b6e-76946c7952b6	737b35e1-d668-4fce-9b6e-76946c7952b6	male	deceased	C71.9			63	489	NA		NA			Included	96881d37-7a3e-4910-b897-19453118fe7c	b337121c-9821-4644-820e-b8c477f6c38a	BRAIN, & CRANIAL NERVES, & SPINAL CORD, (EXCL. VENTRICLE, CEREBELLUM)	CNS-GBM	NEURAL_CREST	CNS	Diffuse glioma	Glioblastoma	'9440/3	Glioblastoma, NOS			85	85-85				SP23639	#615421	b337121c-9821-4644-820e-b8c477f6c38a	d60f54f5-b154-42c4-99fb-cea4e7a33dc7	Brain	tumor	d60f54f5-b154-42c4-99fb-cea4e7a33dc7
612ef912-5a28-4c11-8703-3376f51afef5	Brain Glioblastoma Multiforme - TCGA, US	GBM-US	DO10975	SP23769	SA138622	74139255-a635-4c87-814d-3dd04ed630a8	5e92a5d5-77a4-401a-ade3-e01c95302ec0	612ef912-5a28-4c11-8703-3376f51afef5	Primary tumour - solid tissue	74139255-a635-4c87-814d-3dd04ed630a8	5e92a5d5-77a4-401a-ade3-e01c95302ec0	TCGA-06-0190-01A-01R-1849-01	2013-12-17T00:56:04Z	G17214.TCGA-06-0190-01A-01R-1849-01.2.tar.gz	6283869181	4eddd342a530b2fbd5b30e7bda3998fd	263668a7-e877-4311-9c76-13cf23b1e811	https://cghub.ucsc.edu/cghub/data/analysis/download/263668a7-e877-4311-9c76-13cf23b1e811	BI	ILLUMINA	Illumina HiSeq 2000	7290.WR29124.Solexa-89509.C0FVUACXX120313.P	C0FVU.2	G17214.TCGA-06-0190-01A-01R-1849-01.2.bam	NA	FALSENA	NA		RNA-Seq			yes	fastq	GBM-US::74139255-a635-4c87-814d-3dd04ed630a8	Whitelist	Whitelist	GBM-US::74139255-a635-4c87-814d-3dd04ed630a8	74139255-a635-4c87-814d-3dd04ed630a8	male	deceased	C71.9			62	317	NA		NA			Included	5e92a5d5-77a4-401a-ade3-e01c95302ec0	612ef912-5a28-4c11-8703-3376f51afef5	BRAIN, & CRANIAL NERVES, & SPINAL CORD, (EXCL. VENTRICLE, CEREBELLUM)	CNS-GBM	NEURAL_CREST	CNS	Diffuse glioma	Glioblastoma	'9440/3	Glioblastoma, NOS			NA	SP23769	#615421	612ef912-5a28-4c11-8703-3376f51afef5	c065761d-f775-457f-bda0-4c7c257a701e	Braintumor	c065761d-f775-457f-bda0-4c7c257a701e
56a705f4-fd28-44ff-8a3c-85bc4300c760	Brain Glioblastoma Multiforme - TCGA, US	GBM-US	DO11046	SP23925	SA141293	e17a6048-7a72-42c0-ad3f-97cbff02bc9f	b89a4d5d-ee24-4052-b454-b67169328737	56a705f4-fd28-44ff-8a3c-85bc4300c760	Primary tumour - solid tissue	e17a6048-7a72-42c0-ad3f-97cbff02bc9f	b89a4d5d-ee24-4052-b454-b67169328737	TCGA-19-2629-01A-01R-1850-01	2013-12-17T00:55:03Z	G17640.TCGA-19-2629-01A-01R-1850-01.2.tar.gz	8005247390	993e83cdfb12080c2df290066ca2f684	d0dc97b1-0dd1-40b0-89c5-f7cc748d8ecf	https://cghub.ucsc.edu/cghub/data/analysis/download/d0dc97b1-0dd1-40b0-89c5-f7cc748d8ecf	BI	ILLUMINA	Illumina HiSeq 2000	7290.WR29415.Solexa-90571.C0HTPACXX120405.P	C0HTP.2	G17640.TCGA-19-2629-01A-01R-1850-01.2.bam	NA	FALSENA	NA		RNA-Seq			yes	fastq	GBM-US::e17a6048-7a72-42c0-ad3f-97cbff02bc9f	Whitelist	Whitelist	GBM-US::e17a6048-7a72-42c0-ad3f-97cbff02bc9f	e17a6048-7a72-42c0-ad3f-97cbff02bc9f	male	deceased	C71.9			60	737	NA		NA			Included	b89a4d5d-ee24-4052-b454-b67169328737	56ffaa35-814c-4c0b-b3c6-d4514d34fec2	BRAIN, & CRANIAL NERVES, & SPINAL CORD, (EXCL. VENTRICLE, CEREBELLUM)	CNS-GBM	NEURAL_CREST	CNS	Diffuse glioma	Glioblastoma	'9440/3	Glioblastoma, NOS			85	85-85				SP23925	#615421	56a705f4-fd28-44ff-8a3c-85bc4300c760	56ffaa35-814c-4c0b-b3c6-d4514d34fec2	Brain	tumor	56ffaa35-814c-4c0b-b3c6-d4514d34fec2
28239a0e-9990-49ef-a159-ba63fb078c77	Brain Glioblastoma Multiforme - TCGA, US	GBM-US	DO11148	SP24129	SA144446	0078b0c4-68a9-483b-9aab-61156d263213	f2ffa9ff-86d6-41ef-92e7-7f87e981985b	28239a0e-9990-49ef-a159-ba63fb078c77	Primary tumour - solid tissue	0078b0c4-68a9-483b-9aab-61156d263213	f2ffa9ff-86d6-41ef-92e7-7f87e981985b	TCGA-14-1034-01A-01R-1849-01	2013-12-17T00:53:02Z	G17473.TCGA-14-1034-01A-01R-1849-01.2.tar.gz	4827624481	14a341c7a1dac456e0e9d419551a2c04	98f2d5fa-7957-4a71-bb46-728ff20776ae	https://cghub.ucsc.edu/cghub/data/analysis/download/98f2d5fa-7957-4a71-bb46-728ff20776ae	BI	ILLUMINA	Illumina HiSeq 2000	7290.WR29324.Solexa-91276.C0HTRACXX120406.P	C0HTR.3	G17473.TCGA-14-1034-01A-01R-1849-01.2.bam	NA	FALSENA	NA		RNA-Seq			yes	fastq	GBM-US::0078b0c4-68a9-483b-9aab-61156d263213	Whitelist	Whitelist	GBM-US::0078b0c4-68a9-483b-9aab-61156d263213	0078b0c4-68a9-483b-9aab-61156d263213	female	deceased	C71.9			60	485	NA		NA			Included	f2ffa9ff-86d6-41ef-92e7-7f87e981985b	28239a0e-9990-49ef-a159-ba63fb078c77	BRAIN, & CRANIAL NERVES, & SPINAL CORD, (EXCL. VENTRICLE, CEREBELLUM)	CNS-GBM	NEURAL_CREST	CNS	Diffuse glioma	Glioblastoma	'9440/3	Glioblastoma, NOS			NA	SP24129	#615421	28239a0e-9990-49ef-a159-ba63fb078c77	7cae6c0b-36fe-411b-bbba-093a4c846d84	Braintumor	7cae6c0b-36fe-411b-bbba-093a4c846d84

So each file that contains the information on the extracted tumor as a fileheader similar to the patient ones, but different. In order to match the tumor to the patient it came from, we need the patient reference file to get the patient id, and the tumor reference file (this one) to get the patient id and link the two together. If you're just interested in fixing the analysis parts of the code, don't worry too much about this part. This is what the summary_matching.py file does.

Step 2b

Take the previously built gene-fusion files and add details about exons and location
Genefinder.RNADonorId() will create a new directory called "patient_fusions_with_exons". In here you'll find all the same files from Referenced_patient_data, except now for each fusion in one of those files, you'll see all the exons contained within each gene, as well as their start and stop location.
So this file from Referenced_patient_data

0a6be23a-d5a0-4e95-ada2-a61b2b5d9485.tsv

geneA	geneA orientation	geneB	geneB orientation	chr1	position	chr1 orientation	chr2	position	chr2 orientation
./patient_data/0a6be23a-d5a0-4e95-ada2-a61b2b5d9485.pcawg_consensus_1.6.161116.somatic.sv.bedpe									
ERG	-	TMPRSS2	-	chr21	39867712	+	chr21	42872048	-
ERG	-	TMPRSS2	-	chr21	39883066	-	chr21	42871868	+

will be converted to

./patient_fusions_with_exons/0a6be23a-d5a0-4e95-ada2-a61b2b5d9485.tsv

geneA	geneB		
ERG	TMPRSS2		
ENSG00000184012	TMPRSS2	-	42872048
chr21	-		
1	42880008	42880086	
2	42870046	42870116	
3	42866283	42866505	
4	42861434	42861520	
5	42860321	42860440	
6	42852403	42852529	
7	42851099	42851209	
8	42848504	42848547	
9	42845252	42845423	
10	42843733	42843908	
11	42842575	42842670	
12	42840323	42840465	
13	42839661	42839813	
14	42836478	42838080	
1	42879877	42879909	
2	42870046	42870116	
3	42866283	42866505	
4	42861434	42861520	
5	42860321	42860440	
6	42852403	42852529	
7	42851099	42851209	
8	42848504	42848547	
9	42845252	42845423	
10	42843733	42843908	
11	42842575	42842670	
12	42840323	42840465	
13	42839661	42839813	
14	42837936	42838080	
1	42902912	42903043	
2	42901900	42902145	
3	42870046	42870116	
4	42866283	42866505	
5	42861434	42861520	
6	42860321	42860440	
7	42852403	42852529	
8	42851099	42851209	
9	42848504	42848547	
10	42845252	42845423	
11	42843733	42843908	
12	42842575	42842670	
13	42840323	42840465	
14	42839661	42839813	
1	42878372	42878531	
2	42870046	42870116	
3	42866283	42866505	
			
ENSG00000157554	ERG	-	39867712
chr21	+		
1	39870287	39870407	
2	39817327	39817544	
3	39795332	39795483	
4	39775428	39775631	
5	39774479	39774559	
6	39772496	39772567	
7	39764298	39764366	
8	39763581	39763637	
9	39762917	39762964	
10	39751949	39755845	
1	39956768	39956824	
2	39947586	39947671	
3	39795332	39795483	
4	39775428	39775631	
5	39774479	39774559	
6	39764298	39764366	
7	39763581	39763637	
8	39762917	39762964	
9	39751949	39755845	
1	39956768	39956869	
2	39947586	39947671	
3	39817327	39817544	
4	39795332	39795483	
5	39775428	39775631	
6	39774479	39774559	
7	39764298	39764366	
8	39763581	39763637	
9	39762917	39762964	
10	39751949	39755845	
1	40033582	40033618	
2	39956768	39956869	
3	39947586	39947671	
4	39817327	39817544	
5	39795332	39795483	
6	39775428	39775631	
7	39774479	39774559	
8	39772496	39772567	
9	39764298	39764366	
10	39763581	39763637	
11	39762917	39762964	
12	39751949	39755845	
1	39870287	39870400	
2	39795332	39795483	
3	39775428	39775631	
4	39774479	39774559	
5	39772496	39772567	
6	39764298	39764366	
7	39763581	39763637	
8	39762917	39762964	
9	39755048	39755845	
1	40032446	40032591	
2	39956768	39956869	
3	39947586	39947671	
4	39817327	39817544	
5	39795332	39795483	
6	39775428	39775631	
7	39774479	39774559	
8	39772496	39772567	
9	39764298	39764366	
10	39763581	39763637	
11	39762917	39762964	
12	39755213	39755845	
			
geneA	geneB		
ERG	TMPRSS2		
ENSG00000184012	TMPRSS2	-	42871868
chr21	+		
1	42880008	42880086	
2	42870046	42870116	
3	42866283	42866505	
4	42861434	42861520	
5	42860321	42860440	
6	42852403	42852529	
7	42851099	42851209	
8	42848504	42848547	
9	42845252	42845423	
10	42843733	42843908	
11	42842575	42842670	
12	42840323	42840465	
13	42839661	42839813	
14	42836478	42838080	
1	42879877	42879909	
2	42870046	42870116	
3	42866283	42866505	
4	42861434	42861520	
5	42860321	42860440	
6	42852403	42852529	
7	42851099	42851209	
8	42848504	42848547	
9	42845252	42845423	
10	42843733	42843908	
11	42842575	42842670	
12	42840323	42840465	
13	42839661	42839813	
14	42837936	42838080	
1	42902912	42903043	
2	42901900	42902145	
3	42870046	42870116	
4	42866283	42866505	
5	42861434	42861520	
6	42860321	42860440	
7	42852403	42852529	
8	42851099	42851209	
9	42848504	42848547	
10	42845252	42845423	
11	42843733	42843908	
12	42842575	42842670	
13	42840323	42840465	
14	42839661	42839813	
1	42878372	42878531	
2	42870046	42870116	
3	42866283	42866505	
			
ENSG00000157554	ERG	-	39883066
chr21	-		
1	39870287	39870407	
2	39817327	39817544	
3	39795332	39795483	
4	39775428	39775631	
5	39774479	39774559	
6	39772496	39772567	
7	39764298	39764366	
8	39763581	39763637	
9	39762917	39762964	
10	39751949	39755845	
1	39956768	39956824	
2	39947586	39947671	
3	39795332	39795483	
4	39775428	39775631	
5	39774479	39774559	
6	39764298	39764366	
7	39763581	39763637	
8	39762917	39762964	
9	39751949	39755845	
1	39956768	39956869	
2	39947586	39947671	
3	39817327	39817544	
4	39795332	39795483	
5	39775428	39775631	
6	39774479	39774559	
7	39764298	39764366	
8	39763581	39763637	
9	39762917	39762964	
10	39751949	39755845	
1	40033582	40033618	
2	39956768	39956869	
3	39947586	39947671	
4	39817327	39817544	
5	39795332	39795483	
6	39775428	39775631	
7	39774479	39774559	
8	39772496	39772567	
9	39764298	39764366	
10	39763581	39763637	
11	39762917	39762964	
12	39751949	39755845	
1	39870287	39870400	
2	39795332	39795483	
3	39775428	39775631	
4	39774479	39774559	
5	39772496	39772567	
6	39764298	39764366	
7	39763581	39763637	
8	39762917	39762964	
9	39755048	39755845	
1	40032446	40032591	
2	39956768	39956869	
3	39947586	39947671	
4	39817327	39817544	
5	39795332	39795483	
6	39775428	39775631	
7	39774479	39774559	
8	39772496	39772567	
9	39764298	39764366	
10	39763581	39763637	
11	39762917	39762964	
12	39755213	39755845	

So you can see that the original gene-fusion file had 2 gene fusions, meaning there were a total of 4 genes involved in that file. The new file separates each gene and underneath it, gives all the exon information for each gene.

side note: in Genefinder.py you'll see two functions getExonFreq() and checkExonNumbers(). Those are tests I wrote just to get an idea of what kind of results I was getting for finding exons in that first file. Just stuff like how many occurrences of a specific number I was getting, or how many genes had overlapping numbers, etc. There's no call to those functions in the main operation of the code, so feel free to call them specifically if you're trying to fix that issue. Or if they're not super easy to follow (I didn't comment them cause it was sort of just hacked together pretty quickly), you can write your own inside that file using those arguments.

Step 2c

Next, file_creation.GroupCancers() will create separate directories and put each patient file in the cancer group that they belong in. The reason for this is because when analyzing one patient, we want to look at their RNA-seq values relative to only the other patients with the same cancer type. This is another aspect Dr. Yang can help clarify for you.
After this runs, you'll see a new directory called "CancerType_Groups". Those previous files with exon locations will be sorted into subdirectories, each labeled with the cancer name, within this one. It's sorted this way so that we know which other files to compare the data against.

Step 3

Comparing Exons Scores

Now we're up to the part that you'll be focusing on.

note: this is by far the slowest part. I wrote it for results, not for optimization. You can see for yourself it's O(n 3). However, it's easily adjustable to just run for results on specific cancer types. You don't need to run the script for the entirety of the data set.

filecreation.EvaluatePatientGroup() looks at each exon on each gene for each patient within a cancer group. What it then does is takes that exon's score, as well as the score for that same exon on the same gene, but from all the patients with the same cancer but that don't have that gene fusion.
Basically, think about it like this...

- Every patient (every human, really) has gene "A" (we'll just call it A).  
- Let's say we have 3 lung cancer patients total;

Patient 1 has gene fusions
A-B, D-K and C-F
Patient 2 has gene fusions
C-R, G-D and A-Q
Patient 3 has gene fusions
D-F, Q-G and C-L
  
Now, when we're looking at Patient 1, we look at each individual gene, starting with A. 
Then we first look at every other patient in our group and see which ones of them don't have A involved 
in any gene fusions at all. 
In this case, it's just Patient 3. 
Now Patient 3 still has gene A in their genome and the RNA-seq file contains exon scores for each exon on that gene. 
So we collect the scores of each exon number on gene A for Patient 1 and of each exon on gene A for Patient 3

We convert this array of scores to an array of z scores for each exon (line 208 in file_creation.py)
This is to measure the statistical difference of the exons involved in fusions
versus the exons involved in normally behaving genes.

That last part is is the part you may want to consider changing.
The zscores are used for the T-test, but a T-test isn't really suited for this type of data

At the end of this, you'll have a new directory called "Results". Inside here, you'll see subdirectories labeled by cancer name. Inside each of those, you'll see files with familiar looking file prefixes. Remember the files before that had each individual gene with the exons and their locations? Now those same files have had the associated zscores for each of those exons added to that information. Below is part of one of those files (just the first two individual genes aka, one gene fusion)

ENSG00000083290	ULK2	-	19768956	-
1	19770641,19771249,-0.218217890236			
2	19769049,19769141,-1.00636292769			
3	19768116,19768157,-0.737993368111			
4	19753055,19753087,-0.0133448318556			
5	19752682,19752718,-0.358192041357			
6	19750050,19750223,-0.273878970921			
7	19748617,19748690,-0.088162847883			
8	19746434,19746535,-0.0399628447329			
9	19744802,19744860,0.0203875352017			
10	19741814,19741896,-0.148956553759			
11	19729452,19729499,-0.122528227552			
12	19728407,19728495,0.25224953031			
13	19720062,19720233,-0.933138949632			
14	19713691,19713751,0.293840438991			
15	19708006,19708143,0.089826267105			
16	19705090,19705235,-0.0894836314736			
17	19702808,19702888,0.0209262830808			
18	19700709,19700995,-0.0998118416406			
19	19699408,19699595,-0.581914373963			
20	19698935,19699038,0.0580999938876			
21	19689250,19689399,-0.145679006529			
22	19687010,19687218,0.113141497729			
23	19685202,19685380,0.133191619312			
24	19684299,19684417,0.425414416472			
25	19683785,19683942,0.334668536512			
26	19680894,19681029,0.267610802961			
27	19679622,19679720,-0.298511157063			
28	19674143,19674380,-0.596196475138			
ENSG00000065675	PRKCQ	-	6584565	+
1	6622173,6622263,-0.530281060691			
2	6556980,6557106,-0.632284962063			
3	6552957,6553156,-0.54136434314			
4	6549398,6549458,-0.581810520044			
5	6540358,6540520,-0.571063273502			
6	6539182,6539213,-0.591452355234			
7	6538997,6539082,-0.606197729181			
8	6533645,6533774,-0.66439290649			
9	6527997,6528106,-0.592499311674			
10	6527114,6527231,-0.605737135917			
11	6525402,6525562,-0.599494146932			
12	6520954,6521127,-0.587494011963			
13	6506275,6506366,-0.547822170593			
14	6504265,6504327,-0.568736248817			
15	6498636,6498774,-0.605030192545			
16	6483855,6484043,-0.575022611536			
17	6472772,6472900,-0.566295527583			
18	6469105,6470324,-0.545479126787			

The key to using this data (although, again, you're probably going to end up doing something different than taking zscores, but they're here if you need them) is to use the breakpoint for the gene to split the array of zscores. So for example, in the last gene listed here, you'll see after the gene name and orientation, the number 6584565. That's the location on this gene where it's splitting and recombining with the first gene listed. Now look at the locations of all the exons. You'll notice that exon 1 is above that breakpoint and the rest of them are below that. So the T-test will compare the difference of the set of zscores between those two sides. Again, this is why T-test is super not good. You're comparing 1 datum to 17 other data points. Such unbalance. Much bad. What to do? This is the biggest issue with the data.

Step 4 - Final Results

Evaluate.pvalueStats() and Evaluate.MasterList() use the zscores from these files and a T-test across the breakpoints of the individual genes and sorts by pvalue. pvalueStats() is for each individual cancer type, MasterList will collect all of the data for you and create a list, sorted by best (most likely candidates) first. Essentially how the tests are considered is this: in one gene fusion, you'll have two T-tests done; one on the first gene across its breakpoint (which will have a resulting pvalue) and the second on the other gene, also with a resulting pvalue. Because only one gene in a fusion is all that's required to be a driver (for instance, if gene A is a driver, you could have fusions A-B, A-L, A-P etc all being oncogenic) we take the best p value of the two and assign the entire fusion that best score. My current top 10 list after running this script (and waiting a long time for it to complete) looks like this.

#columns: first_gene, second_gene, best_pvalue_of_the_two, cancer type
ALK	TRPC4AP	0.0001	LUAD
SLC7A6	PRKCA	0.00016	LUSC
KDELC2	SIK2	0.00016	LUSC
GPR116	MIOS	0.00019	LUAD
RAP1GDS1	NPNT	0.00021	LUAD
GANC	PDE8A	0.00022	LUAD
FARP2	ABCB4	0.00022	HNSC
CAPRIN2	PPM1L	0.00024	HNSC
MAP3K3	SCN4A	0.00025	HNSC
C14orf39	PCLO	0.00028	LUSC

That's about as much as I think I can include to get you started. Feel free to email me at collinol@uchicago.edu for questions or clarifications. Like I said, I suspect your main goal that Dr. Yang will task you with will be to develop a better method of scoring the fusions. If you need help reconfiguring the files or maybe getting some data somewhere along the process that I didn't include, let me know. I should be able to find it for you so you don't have to waste time looking through csv columns.

Additionally, for better statistical tests, I suspect that will be the major focus of your work. Deciding which tests you think would work best, given the nature of the data. My research ended before I could continue with that, so I don't have all the answers. However, before I stopped working on it, I was looking at changing the t-test over to variance analysis like you mentioned. Specifically, ANOVA (analysis of variance) test. There's some good documentation on it here http://www.statisticshowto.com/probability-and-statistics/hypothesis-testing/anova/#HowToRun. It most likely won't be a perfect fit, but there's already python libraries for it https://plot.ly/python/anova/. So it should be relatively quick to implement and see what results you get.

Beyond pre-built stats packages, there are known driver fusions (ERG-TMPRSS2, BCR-ABL, ALK-(with lots of others), etc). I'm not sure how familiar you are with machine learning techniques, but if you have the time to invest into it, it could be a good approach to collect the data from the current scripts regarding only the exon values for known gene fusions, split that information into a training and testing group, and then see if you can develop a model to accurately classify the test data. I'd start with binary classification (known fusions, unknown fusions) and then get more specific as to what the actual gene fusion is.

Good luck!

Collin Olander

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