Tues. Aug. 6, 2019 Salmon + Sea lice BS data analysis

(base) ostrich:TRIM_5bp strigg$ /Users/strigg/anaconda3/bin/TrimGalore-0.6.0/trim_galore \
> --output_dir /Users/strigg/Desktop/Salmo_Calig/TRIM_5bp \
> --fastqc_args \
> "--outdir /Users/strigg/Desktop/Salmo_Calig/TRIM_5bp/FASTQC" \
> --clip_R1 5 \
> --clip_R2 5 \
> --paired \
> --path_to_cutadapt /Users/strigg/anaconda3/bin/cutadapt \
> /Users/strigg/Desktop/Salmo_Calig/16C_26psu_1_S13_L001_R1_001.fastq.gz \
> /Users/strigg/Desktop/Salmo_Calig/16C_26psu_1_S13_L001_R2_001.fastq.gz 
Multicore support not enabled. Proceeding with single-core trimming.
Path to Cutadapt set as: '/Users/strigg/anaconda3/bin/cutadapt' (user defined)
Cutadapt seems to be working fine (tested command '/Users/strigg/anaconda3/bin/cutadapt --version')
No quality encoding type selected. Assuming that the data provided uses Sanger encoded Phred scores (default)

Output directory /Users/strigg/Desktop/Salmo_Calig/TRIM_5bp/ doesn't exist, creating it for you...

Output will be written into the directory: /Users/strigg/Desktop/Salmo_Calig/TRIM_5bp/


AUTO-DETECTING ADAPTER TYPE
===========================
Attempting to auto-detect adapter type from the first 1 million sequences of the first file (>> /Users/strigg/Desktop/Salmo_Calig/16C_26psu_1_S13_L001_R1_001.fastq.gz <<)

Found perfect matches for the following adapter sequences:
Adapter type	Count	Sequence	Sequences analysed	Percentage
Illumina	538094	AGATCGGAAGAGC	1000000	53.81
Nextera	0	CTGTCTCTTATA	1000000	0.00
smallRNA	0	TGGAATTCTCGG	1000000	0.00
Using Illumina adapter for trimming (count: 538094). Second best hit was Nextera (count: 0)

Writing report to '/Users/strigg/Desktop/Salmo_Calig/TRIM_5bp/16C_26psu_1_S13_L001_R1_001.fastq.gz_trimming_report.txt'

SUMMARISING RUN PARAMETERS
==========================
Input filename: /Users/strigg/Desktop/Salmo_Calig/16C_26psu_1_S13_L001_R1_001.fastq.gz
Trimming mode: paired-end
Trim Galore version: 0.6.0
Cutadapt version: 2.4
Number of cores used for trimming: 1
Quality Phred score cutoff: 20
Quality encoding type selected: ASCII+33
Adapter sequence: 'AGATCGGAAGAGC' (Illumina TruSeq, Sanger iPCR; auto-detected)
Maximum trimming error rate: 0.1 (default)
Minimum required adapter overlap (stringency): 1 bp
Minimum required sequence length for both reads before a sequence pair gets removed: 20 bp
All Read 1 sequences will be trimmed by 5 bp from their 5' end to avoid poor qualities or biases
All Read 2 sequences will be trimmed by 5 bp from their 5' end to avoid poor qualities or biases (e.g. M-bias for BS-Seq applications)
Running FastQC on the data once trimming has completed
Running FastQC with the following extra arguments: '--outdir /Users/strigg/Desktop/Salmo_Calig/TRIM_5bp/FASTQC'
Output file(s) will be GZIP compressed

Writing final adapter and quality trimmed output to 16C_26psu_1_S13_L001_R1_001_trimmed.fq.gz


  >>> Now performing quality (cutoff '-q 20') and adapter trimming in a single pass for the adapter sequence: 'AGATCGGAAGAGC' from file /Users/strigg/Desktop/Salmo_Calig/16C_26psu_1_S13_L001_R1_001.fastq.gz <<< 
10000000 sequences processed
20000000 sequences processed
This is cutadapt 2.4 with Python 3.6.8
Command line parameters: -j 1 -e 0.1 -q 20 -O 1 -a AGATCGGAAGAGC /Users/strigg/Desktop/Salmo_Calig/16C_26psu_1_S13_L001_R1_001.fastq.gz
Processing reads on 1 core in single-end mode ...
Finished in 852.63 s (38 us/read; 1.56 M reads/minute).

=== Summary ===

Total reads processed:              22,159,935
Reads with adapters:                17,366,785 (78.4%)
Reads written (passing filters):    22,159,935 (100.0%)

Total basepairs processed: 3,323,990,250 bp
Quality-trimmed:              20,636,466 bp (0.6%)
Total written (filtered):  2,599,584,885 bp (78.2%)

=== Adapter 1 ===

Sequence: AGATCGGAAGAGC; Type: regular 3'; Length: 13; Trimmed: 17366785 times.

No. of allowed errors:
0-9 bp: 0; 10-13 bp: 1

Bases preceding removed adapters:
  A: 29.9%
  C: 14.0%
  G: 20.5%
  T: 35.6%
  none/other: 0.0%

Overview of removed sequences
length	count	expect	max.err	error counts
1	2904827	5539983.8	0	2904827
2	423079	1384995.9	0	423079
3	231866	346249.0	0	231866
4	143449	86562.2	0	143449
5	117570	21640.6	0	117570
6	115529	5410.1	0	115529
7	96119	1352.5	0	96119
8	119364	338.1	0	119364
9	118406	84.5	0	116886 1520
10	112051	21.1	1	106032 6019
11	122987	5.3	1	113734 9253
12	119477	1.3	1	111842 7635
13	120023	0.3	1	112246 7777
14	122963	0.3	1	114085 8878
15	127528	0.3	1	119209 8319
16	126445	0.3	1	117002 9443
17	152761	0.3	1	140488 12273
18	129037	0.3	1	121213 7824
19	112889	0.3	1	105773 7116
20	133616	0.3	1	125004 8612
21	146516	0.3	1	136291 10225
22	126419	0.3	1	119045 7374
23	145391	0.3	1	135716 9675
24	136894	0.3	1	125670 11224
25	145117	0.3	1	135313 9804
26	134897	0.3	1	126717 8180
27	148757	0.3	1	137734 11023
28	145635	0.3	1	135492 10143
29	153188	0.3	1	140948 12240
30	163373	0.3	1	154030 9343
31	130689	0.3	1	122129 8560
32	157390	0.3	1	147156 10234
33	145226	0.3	1	135269 9957
34	158524	0.3	1	147826 10698
35	161582	0.3	1	148734 12848
36	173098	0.3	1	162838 10260
37	141921	0.3	1	133053 8868
38	164933	0.3	1	154468 10465
39	167185	0.3	1	155913 11272
40	175799	0.3	1	163207 12592
41	197239	0.3	1	176427 20812
42	165247	0.3	1	150517 14730
43	270149	0.3	1	254538 15611
44	85600	0.3	1	76961 8639
45	197628	0.3	1	175697 21931
46	723044	0.3	1	688287 34757
47	254889	0.3	1	240372 14517
48	114855	0.3	1	101708 13147
49	409546	0.3	1	390013 19533
50	125438	0.3	1	116192 9246
51	110069	0.3	1	99667 10402
52	381945	0.3	1	364475 17470
53	196688	0.3	1	188239 8449
54	63003	0.3	1	58174 4829
55	106109	0.3	1	101117 4992
56	55521	0.3	1	50154 5367
57	169184	0.3	1	161794 7390
58	35182	0.3	1	32212 2970
59	44244	0.3	1	39185 5059
60	173147	0.3	1	165425 7722
61	86440	0.3	1	82402 4038
62	40559	0.3	1	36479 4080
63	130914	0.3	1	123404 7510
64	122315	0.3	1	116337 5978
65	65805	0.3	1	60428 5377
66	148115	0.3	1	139100 9015
67	175930	0.3	1	163369 12561
68	293688	0.3	1	278052 15636
69	189726	0.3	1	179114 10612
70	75474	0.3	1	69457 6017
71	27345	0.3	1	23952 3393
72	97969	0.3	1	91968 6001
73	138145	0.3	1	130111 8034
74	143624	0.3	1	135279 8345
75	142674	0.3	1	134066 8608
76	141682	0.3	1	133308 8374
77	141205	0.3	1	132947 8258
78	133036	0.3	1	124966 8070
79	131630	0.3	1	123633 7997
80	128859	0.3	1	121116 7743
81	126119	0.3	1	118667 7452
82	123492	0.3	1	116262 7230
83	118847	0.3	1	111615 7232
84	115583	0.3	1	108642 6941
85	111409	0.3	1	104817 6592
86	107099	0.3	1	100545 6554
87	102731	0.3	1	96477 6254
88	101527	0.3	1	95245 6282
89	95383	0.3	1	89600 5783
90	95037	0.3	1	89230 5807
91	89836	0.3	1	84395 5441
92	84977	0.3	1	79865 5112
93	79479	0.3	1	74762 4717
94	76672	0.3	1	72010 4662
95	70676	0.3	1	66313 4363
96	66886	0.3	1	62766 4120
97	63105	0.3	1	59150 3955
98	58813	0.3	1	55101 3712
99	56801	0.3	1	52921 3880
100	51251	0.3	1	47973 3278
101	48131	0.3	1	44969 3162
102	43975	0.3	1	41134 2841
103	40749	0.3	1	38132 2617
104	37602	0.3	1	35059 2543
105	33601	0.3	1	31318 2283
106	30687	0.3	1	28622 2065
107	27550	0.3	1	25698 1852
108	25919	0.3	1	24135 1784
109	23271	0.3	1	21584 1687
110	20524	0.3	1	19077 1447
111	18563	0.3	1	17263 1300
112	16204	0.3	1	15036 1168
113	14110	0.3	1	13017 1093
114	12321	0.3	1	11394 927
115	10629	0.3	1	9884 745
116	9483	0.3	1	8756 727
117	8057	0.3	1	7440 617
118	7067	0.3	1	6491 576
119	5777	0.3	1	5264 513
120	4917	0.3	1	4511 406
121	4181	0.3	1	3800 381
122	3506	0.3	1	3174 332
123	2912	0.3	1	2640 272
124	2546	0.3	1	2297 249
125	2101	0.3	1	1902 199
126	1717	0.3	1	1548 169
127	1536	0.3	1	1361 175
128	1201	0.3	1	1095 106
129	1012	0.3	1	912 100
130	739	0.3	1	668 71
131	638	0.3	1	572 66
132	553	0.3	1	500 53
133	556	0.3	1	500 56
134	665	0.3	1	607 58
135	362	0.3	1	324 38
136	207	0.3	1	176 31
137	143	0.3	1	128 15
138	99	0.3	1	86 13
139	84	0.3	1	72 12
140	58	0.3	1	50 8
141	42	0.3	1	38 4
142	37	0.3	1	29 8
143	24	0.3	1	22 2
144	25	0.3	1	17 8
145	26	0.3	1	22 4
146	29	0.3	1	21 8
147	85	0.3	1	77 8
148	64	0.3	1	59 5
149	40	0.3	1	35 5
150	355	0.3	1	318 37

RUN STATISTICS FOR INPUT FILE: /Users/strigg/Desktop/Salmo_Calig/16C_26psu_1_S13_L001_R1_001.fastq.gz
=============================================
22159935 sequences processed in total
The length threshold of paired-end sequences gets evaluated later on (in the validation step)

Writing report to '/Users/strigg/Desktop/Salmo_Calig/TRIM_5bp/16C_26psu_1_S13_L001_R2_001.fastq.gz_trimming_report.txt'

SUMMARISING RUN PARAMETERS
==========================
Input filename: /Users/strigg/Desktop/Salmo_Calig/16C_26psu_1_S13_L001_R2_001.fastq.gz
Trimming mode: paired-end
Trim Galore version: 0.6.0
Cutadapt version: 2.4
Number of cores used for trimming: 1
Quality Phred score cutoff: 20
Quality encoding type selected: ASCII+33
Adapter sequence: 'AGATCGGAAGAGC' (Illumina TruSeq, Sanger iPCR; auto-detected)
Maximum trimming error rate: 0.1 (default)
Minimum required adapter overlap (stringency): 1 bp
Minimum required sequence length for both reads before a sequence pair gets removed: 20 bp
All Read 1 sequences will be trimmed by 5 bp from their 5' end to avoid poor qualities or biases
All Read 2 sequences will be trimmed by 5 bp from their 5' end to avoid poor qualities or biases (e.g. M-bias for BS-Seq applications)
Running FastQC on the data once trimming has completed
Running FastQC with the following extra arguments: '--outdir /Users/strigg/Desktop/Salmo_Calig/TRIM_5bp/FASTQC'
Output file(s) will be GZIP compressed

Writing final adapter and quality trimmed output to 16C_26psu_1_S13_L001_R2_001_trimmed.fq.gz


  >>> Now performing quality (cutoff '-q 20') and adapter trimming in a single pass for the adapter sequence: 'AGATCGGAAGAGC' from file /Users/strigg/Desktop/Salmo_Calig/16C_26psu_1_S13_L001_R2_001.fastq.gz <<< 
10000000 sequences processed
20000000 sequences processed
This is cutadapt 2.4 with Python 3.6.8
Command line parameters: -j 1 -e 0.1 -q 20 -O 1 -a AGATCGGAAGAGC /Users/strigg/Desktop/Salmo_Calig/16C_26psu_1_S13_L001_R2_001.fastq.gz
Processing reads on 1 core in single-end mode ...
Finished in 904.27 s (41 us/read; 1.47 M reads/minute).

=== Summary ===

Total reads processed:              22,159,935
Reads with adapters:                17,263,070 (77.9%)
Reads written (passing filters):    22,159,935 (100.0%)

Total basepairs processed: 3,323,990,250 bp
Quality-trimmed:              16,760,179 bp (0.5%)
Total written (filtered):  2,603,569,143 bp (78.3%)

=== Adapter 1 ===

Sequence: AGATCGGAAGAGC; Type: regular 3'; Length: 13; Trimmed: 17263070 times.

No. of allowed errors:
0-9 bp: 0; 10-13 bp: 1

Bases preceding removed adapters:
  A: 28.6%
  C: 12.4%
  G: 23.7%
  T: 35.3%
  none/other: 0.0%

Overview of removed sequences
length	count	expect	max.err	error counts
1	2980698	5539983.8	0	2980698
2	434290	1384995.9	0	434290
3	244115	346249.0	0	244115
4	143736	86562.2	0	143736
5	130207	21640.6	0	130207
6	112454	5410.1	0	112454
7	91383	1352.5	0	91383
8	130413	338.1	0	130413
9	102897	84.5	0	102495 402
10	113241	21.1	1	107719 5522
11	119188	5.3	1	111572 7616
12	120663	1.3	1	112735 7928
13	133942	0.3	1	125965 7977
14	126547	0.3	1	117452 9095
15	102830	0.3	1	96138 6692
16	149163	0.3	1	139761 9402
17	119889	0.3	1	111631 8258
18	113552	0.3	1	106594 6958
19	144826	0.3	1	135202 9624
20	135016	0.3	1	126722 8294
21	118040	0.3	1	110327 7713
22	135122	0.3	1	126795 8327
23	139623	0.3	1	130868 8755
24	139327	0.3	1	130312 9015
25	156486	0.3	1	147346 9140
26	131367	0.3	1	122873 8494
27	144343	0.3	1	134126 10217
28	135732	0.3	1	128445 7287
29	144460	0.3	1	135341 9119
30	143460	0.3	1	135475 7985
31	154935	0.3	1	145507 9428
32	140211	0.3	1	132501 7710
33	154714	0.3	1	145023 9691
34	197197	0.3	1	185506 11691
35	114069	0.3	1	107701 6368
36	154713	0.3	1	145095 9618
37	166945	0.3	1	157027 9918
38	155394	0.3	1	147358 8036
39	151879	0.3	1	143914 7965
40	168764	0.3	1	158797 9967
41	162964	0.3	1	154146 8818
42	166211	0.3	1	157158 9053
43	152395	0.3	1	144005 8390
44	174656	0.3	1	164812 9844
45	186871	0.3	1	176811 10060
46	135036	0.3	1	127666 7370
47	168636	0.3	1	159853 8783
48	166288	0.3	1	157802 8486
49	174530	0.3	1	165351 9179
50	164769	0.3	1	155907 8862
51	207765	0.3	1	197011 10754
52	147151	0.3	1	139795 7356
53	177051	0.3	1	168817 8234
54	133625	0.3	1	126900 6725
55	174567	0.3	1	166338 8229
56	178950	0.3	1	169886 9064
57	168945	0.3	1	161037 7908
58	161931	0.3	1	154618 7313
59	165789	0.3	1	157874 7915
60	171736	0.3	1	163328 8408
61	175826	0.3	1	167046 8780
62	192123	0.3	1	182534 9589
63	252743	0.3	1	241513 11230
64	207112	0.3	1	199124 7988
65	135660	0.3	1	129354 6306
66	145866	0.3	1	139110 6756
67	145748	0.3	1	138731 7017
68	146161	0.3	1	139522 6639
69	144487	0.3	1	137768 6719
70	154292	0.3	1	147160 7132
71	158353	0.3	1	151327 7026
72	150227	0.3	1	143181 7046
73	157376	0.3	1	150704 6672
74	144388	0.3	1	138042 6346
75	137234	0.3	1	131094 6140
76	133825	0.3	1	127901 5924
77	133664	0.3	1	127640 6024
78	127632	0.3	1	121889 5743
79	122934	0.3	1	117397 5537
80	122791	0.3	1	117317 5474
81	119094	0.3	1	113976 5118
82	115048	0.3	1	109953 5095
83	110957	0.3	1	106119 4838
84	107760	0.3	1	102959 4801
85	105919	0.3	1	100973 4946
86	104575	0.3	1	99936 4639
87	105119	0.3	1	100537 4582
88	102473	0.3	1	97892 4581
89	90861	0.3	1	86820 4041
90	87725	0.3	1	83791 3934
91	82796	0.3	1	79052 3744
92	76768	0.3	1	73386 3382
93	74497	0.3	1	71224 3273
94	71670	0.3	1	68502 3168
95	67300	0.3	1	64357 2943
96	63751	0.3	1	61088 2663
97	58781	0.3	1	56250 2531
98	54871	0.3	1	52462 2409
99	53000	0.3	1	50773 2227
100	48407	0.3	1	46381 2026
101	45093	0.3	1	43177 1916
102	40752	0.3	1	38996 1756
103	37584	0.3	1	35906 1678
104	35749	0.3	1	34226 1523
105	31850	0.3	1	30464 1386
106	29282	0.3	1	28064 1218
107	26084	0.3	1	24956 1128
108	23726	0.3	1	22669 1057
109	21646	0.3	1	20704 942
110	19323	0.3	1	18515 808
111	18235	0.3	1	17436 799
112	15797	0.3	1	15095 702
113	13713	0.3	1	13089 624
114	11968	0.3	1	11416 552
115	10124	0.3	1	9656 468
116	8777	0.3	1	8354 423
117	7339	0.3	1	6995 344
118	6452	0.3	1	6137 315
119	5422	0.3	1	5101 321
120	4499	0.3	1	4281 218
121	3883	0.3	1	3680 203
122	3313	0.3	1	3156 157
123	2807	0.3	1	2669 138
124	2400	0.3	1	2285 115
125	2022	0.3	1	1907 115
126	1650	0.3	1	1555 95
127	1434	0.3	1	1346 88
128	1129	0.3	1	1055 74
129	969	0.3	1	902 67
130	682	0.3	1	644 38
131	587	0.3	1	552 35
132	523	0.3	1	500 23
133	524	0.3	1	494 30
134	633	0.3	1	604 29
135	337	0.3	1	318 19
136	197	0.3	1	186 11
137	147	0.3	1	133 14
138	92	0.3	1	88 4
139	79	0.3	1	71 8
140	57	0.3	1	52 5
141	37	0.3	1	35 2
142	37	0.3	1	35 2
143	25	0.3	1	20 5
144	20	0.3	1	20
145	26	0.3	1	25 1
146	24	0.3	1	23 1
147	85	0.3	1	85
148	61	0.3	1	59 2
149	39	0.3	1	36 3
150	325	0.3	1	310 15

RUN STATISTICS FOR INPUT FILE: /Users/strigg/Desktop/Salmo_Calig/16C_26psu_1_S13_L001_R2_001.fastq.gz
=============================================
22159935 sequences processed in total
The length threshold of paired-end sequences gets evaluated later on (in the validation step)

Validate paired-end files 16C_26psu_1_S13_L001_R1_001_trimmed.fq.gz and 16C_26psu_1_S13_L001_R2_001_trimmed.fq.gz
file_1: 16C_26psu_1_S13_L001_R1_001_trimmed.fq.gz, file_2: 16C_26psu_1_S13_L001_R2_001_trimmed.fq.gz


>>>>> Now validing the length of the 2 paired-end infiles: 16C_26psu_1_S13_L001_R1_001_trimmed.fq.gz and 16C_26psu_1_S13_L001_R2_001_trimmed.fq.gz <<<<<
Writing validated paired-end read 1 reads to 16C_26psu_1_S13_L001_R1_001_val_1.fq.gz
Writing validated paired-end read 2 reads to 16C_26psu_1_S13_L001_R2_001_val_2.fq.gz

Total number of sequences analysed: 22159935

Number of sequence pairs removed because at least one read was shorter than the length cutoff (20 bp): 42248 (0.19%)


  >>> Now running FastQC on the validated data 16C_26psu_1_S13_L001_R1_001_val_1.fq.gz<<<

Specified output directory '/Users/strigg/Desktop/Salmo_Calig/TRIM_5bp/FASTQC' does not exist

  >>> Now running FastQC on the validated data 16C_26psu_1_S13_L001_R2_001_val_2.fq.gz<<<

Specified output directory '/Users/strigg/Desktop/Salmo_Calig/TRIM_5bp/FASTQC' does not exist
Deleting both intermediate output files 16C_26psu_1_S13_L001_R1_001_trimmed.fq.gz and 16C_26psu_1_S13_L001_R2_001_trimmed.fq.gz

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