Bowtie seems to be working fine (tested command '/gscratch/srlab/programs/bowtie2-2.3.4.1-linux-x86_64/bowtie2 --version' [2.3.4]) Output format is BAM (default) Alignments will be written out in BAM format. Samtools found here: '/gscratch/srlab/programs/samtools-1.9/samtools' Reference genome folder provided is /gscratch/srlab/sam/data/C_gigas/genomes/ (absolute path is '/gscratch/srlab/sam/data/C_gigas/genomes/)' Processing sequences up to read no. 5000000 from the input file Attention: using more than 4 cores per alignment thread has been reported to have diminishing returns. If possible try to limit -p to a value of 4 Input files to be analysed (in current folder '/gscratch/scrubbed/samwhite/outputs/20190222_cgigas_se_bismark/subset_5000000'): /gscratch/scrubbed/samwhite/data/C_gigas/BSseq/cgig_bsseq_all_R1.fastq.gz Library is assumed to be strand-specific (directional), alignments to strands complementary to the original top or bottom strands will be ignored (i.e. not performed!) Setting parallelization to single-threaded (default) Current working directory is: /gscratch/scrubbed/samwhite/outputs/20190222_cgigas_se_bismark/subset_5000000 Now reading in and storing sequence information of the genome specified in: /gscratch/srlab/sam/data/C_gigas/genomes/ Single-core mode: setting pid to 1 Single-end alignments will be performed ======================================= Input file is in FastQ format Processing reads up to sequence no. 5000000 from /gscratch/scrubbed/samwhite/data/C_gigas/BSseq/cgig_bsseq_all_R1.fastq.gz Writing a C -> T converted version of the input file cgig_bsseq_all_R1.fastq.gz to cgig_bsseq_all_R1.fastq.gz_C_to_T.fastq Created C -> T converted version of the FastQ file cgig_bsseq_all_R1.fastq.gz (5000001 sequences in total) Input file is cgig_bsseq_all_R1.fastq.gz_C_to_T.fastq (FastQ) Now running 2 instances of Bowtie 2 against the bisulfite genome of /gscratch/srlab/sam/data/C_gigas/genomes/ with the specified options: -q --score-min L,0,-0.6 -p 28 --reorder --ignore-quals Now starting the Bowtie 2 aligner for CTreadCTgenome (reading in sequences from cgig_bsseq_all_R1.fastq.gz_C_to_T.fastq with options -q --score-min L,0,-0.6 -p 28 --reorder --ignore-quals --norc) Using Bowtie 2 index: /gscratch/srlab/sam/data/C_gigas/genomes/Bisulfite_Genome/CT_conversion/BS_CT Found first alignment: HWI-D00743:54:C8MOMACXX:1:1101:1599:2153_1:N:0:TAGCTT 4 * 0 0 * * 0 0 TNGGGGTAGGTTGAATTTGTATTTTTTATGTAGTTTTTTGAGAAAGATTGA C#1AD;2@;D>> Writing bisulfite mapping results to cgig_bsseq_all_R1_bismark_bt2.bam <<< Reading in the sequence file /gscratch/scrubbed/samwhite/data/C_gigas/BSseq/cgig_bsseq_all_R1.fastq.gz Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1101:1773:34563_1:N:0:TAGCTT scaffold33742 2 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1102:3577:18659_1:N:0:TAGCTT C23010 2001 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1103:12835:3587_1:N:0:TAGCTT C23010 2002 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1104:10421:24530_1:N:0:TAGCTT scaffold38030 1 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1104:18665:62350_1:N:0:TAGCTT C34600 19430 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1104:2988:100874_1:N:0:TAGCTT scaffold811 2 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1105:13327:43238_1:N:0:TAGCTT C28016 2 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1105:12755:63320_1:N:0:TAGCTT C32222 11023 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1105:20213:64837_1:N:0:TAGCTT scaffold845 3217 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1106:12588:36512_1:N:0:TAGCTT scaffold838 1 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1108:15893:2479_1:N:0:TAGCTT C35992 1 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1108:10332:16567_1:N:0:TAGCTT C15208 2 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1108:19128:57495_1:N:0:TAGCTT C14656 214 Processed 1000000 sequences so far Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1110:6194:34264_1:N:0:TAGCTT scaffold38856 2 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1110:1976:74862_1:N:0:TAGCTT C15654 2 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1110:17016:83695_1:N:0:TAGCTT C15162 2 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1110:8021:96188_1:N:0:TAGCTT scaffold1895 1 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1111:7845:68788_1:N:0:TAGCTT C32222 11023 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1111:20543:93295_1:N:0:TAGCTT scaffold38030 1 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1111:5799:96032_1:N:0:TAGCTT scaffold26586 3991 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1112:5325:37641_1:N:0:TAGCTT C32222 11023 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1112:3805:88811_1:N:0:TAGCTT C33332 13914 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1112:7692:91397_1:N:0:TAGCTT scaffold33742 2 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1114:18516:50545_1:N:0:TAGCTT C23010 2002 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1114:21097:60128_1:N:0:TAGCTT scaffold39464 72101 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1114:21061:89547_1:N:0:TAGCTT scaffold42790 2 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1115:9662:19845_1:N:0:TAGCTT C14724 214 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1115:14174:28027_1:N:0:TAGCTT scaffold681 2 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1115:9463:37538_1:N:0:TAGCTT scaffold25910 1 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1115:10253:44431_1:N:0:TAGCTT scaffold33902 2 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1115:8202:63048_1:N:0:TAGCTT scaffold33742 2 Processed 2000000 sequences so far Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1116:1971:28732_1:N:0:TAGCTT C20886 1174 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1201:11745:34366_1:N:0:TAGCTT C32222 11023 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1202:4837:35030_1:N:0:TAGCTT C32222 11025 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1202:4675:50032_1:N:0:TAGCTT C34418 18492 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1202:20402:88707_1:N:0:TAGCTT scaffold1359 1 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1203:12182:58735_1:N:0:TAGCTT scaffold33876 15779 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1203:10076:68867_1:N:0:TAGCTT scaffold221 207838 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1203:19553:84916_1:N:0:TAGCTT C31482 1 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1203:7598:91082_1:N:0:TAGCTT C22634 1872 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1204:7258:18231_1:N:0:TAGCTT scaffold36492 35558 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1204:4692:30668_1:N:0:TAGCTT scaffold998 153228 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1204:10882:82970_1:N:0:TAGCTT C22904 1957 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1205:2960:31454_1:N:0:TAGCTT C21854 1 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1205:9982:65799_1:N:0:TAGCTT C16622 349 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1206:11090:25238_1:N:0:TAGCTT C26688 4076 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1206:14430:79019_1:N:0:TAGCTT C23010 2001 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1206:1401:88696_1:N:0:TAGCTT C28852 5906 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1207:9715:8062_1:N:0:TAGCTT C30430 2 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1207:1404:75751_1:N:0:TAGCTT C34600 19430 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1207:8439:81929_1:N:0:TAGCTT scaffold41902 137599 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1207:8338:84034_1:N:0:TAGCTT scaffold67 2 Processed 3000000 sequences so far Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1208:5613:74086_1:N:0:TAGCTT C23010 2001 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1208:12652:82092_1:N:0:TAGCTT C14162 189 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1208:6354:90848_1:N:0:TAGCTT scaffold24070 2508 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1209:2381:44199_1:N:0:TAGCTT C32222 11023 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1209:12796:61059_1:N:0:TAGCTT C15646 266 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1209:1757:76485_1:N:0:TAGCTT C21542 1416 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1209:2728:92564_1:N:0:TAGCTT C32222 11023 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1210:7762:19095_1:N:0:TAGCTT scaffold38366 1 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1210:1459:21708_1:N:0:TAGCTT C17474 2 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1210:12973:39899_1:N:0:TAGCTT C14468 210 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1210:13942:42914_1:N:0:TAGCTT scaffold650 2 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1211:14032:18993_1:N:0:TAGCTT C13236 163 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1211:14913:22666_1:N:0:TAGCTT C16206 2 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1211:6654:67843_1:N:0:TAGCTT scaffold24958 2956 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1211:16110:91525_1:N:0:TAGCTT C32222 11023 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1211:11850:100859_1:N:0:TAGCTT C28430 5506 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1212:16290:18282_1:N:0:TAGCTT scaffold620 3381 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1212:15139:34329_1:N:0:TAGCTT scaffold38856 1 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1212:1416:41915_1:N:0:TAGCTT C32222 11023 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1212:10030:58854_1:N:0:TAGCTT C21854 1 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1212:9970:86696_1:N:0:TAGCTT C13136 1 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1214:15798:11407_1:N:0:TAGCTT scaffold52 3659 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1214:3048:75820_1:N:0:TAGCTT C14656 214 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1215:10174:11135_1:N:0:TAGCTT scaffold1866 525892 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1215:6071:11763_1:N:0:TAGCTT C25616 1 Processed 4000000 sequences so far Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1215:11760:44570_1:N:0:TAGCTT C29764 1 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1215:19336:76957_1:N:0:TAGCTT C24604 1 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1301:19957:51952_1:N:0:TAGCTT scaffold806 114376 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1302:17969:67899_1:N:0:TAGCTT scaffold24958 2956 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1303:1389:25589_1:N:0:TAGCTT C23732 2353 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1303:14402:73643_1:N:0:TAGCTT C23648 2 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1303:17477:95517_1:N:0:TAGCTT scaffold39244 2 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1304:16972:15425_1:N:0:TAGCTT C26492 1 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1304:1698:31473_1:N:0:TAGCTT C14220 2 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1304:2158:66186_1:N:0:TAGCTT C32222 11024 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1306:14437:21907_1:N:0:TAGCTT scaffold42790 1 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1306:20243:23399_1:N:0:TAGCTT scaffold33876 15778 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1306:1150:83570_1:N:0:TAGCTT C20294 983 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1307:15853:25438_1:N:0:TAGCTT scaffold428 2 Chromosomal sequence could not be extracted for HWI-D00743:54:C8MOMACXX:1:1307:13759:27470_1:N:0:TAGCTT scaffold38136 2 5000000 reads; of these: 5000000 (100.00%) were unpaired; of these: 2260986 (45.22%) aligned 0 times 1268229 (25.36%) aligned exactly 1 time 1470785 (29.42%) aligned >1 times 54.78% overall alignment rate 5000000 reads; of these: 5000000 (100.00%) were unpaired; of these: 2257548 (45.15%) aligned 0 times 1269474 (25.39%) aligned exactly 1 time 1472978 (29.46%) aligned >1 times 54.85% overall alignment rate Processed 5000000 sequences so far Successfully deleted the temporary file cgig_bsseq_all_R1.fastq.gz_C_to_T.fastq Final Alignment report ====================== Sequences analysed in total: 5000000 Number of alignments with a unique best hit from the different alignments: 2747063 Mapping efficiency: 54.9% Final Cytosine Methylation Report ================================= Total number of C's analysed: 23218150 Total methylated C's in CpG context: 924940 Total methylated C's in CHG context: 145279 Total methylated C's in CHH context: 276054 Total methylated C's in Unknown context: 4421 Total unmethylated C's in CpG context: 2629317 Total unmethylated C's in CHG context: 4788146 Total unmethylated C's in CHH context: 14454414 Total unmethylated C's in Unknown context: 38351 C methylated in CpG context: 26.0% C methylated in CHG context: 2.9% C methylated in CHH context: 1.9% C methylated in Unknown context (CN or CHN): 10.3% Bismark completed in 0d 0h 25m 23s ==================== Bismark run complete ====================