# Sample Submission – Geoduck Gonad for RNA-seq

Prepared two pools of geoduck RNA for RNA-seq (Illumina HiSeq2500, 100bp, PE) with GENEWIZ, Inc.

I pooled a set of female and a set of male RNAs that had been selected by Steven based on the Bioanalyzer results from Friday.

The female RNA pool used 210ng of each sample, with the exception being sample #08. This sample used 630ng. The reason for this was due to the fact that there weren’t any other female samples to use from this developmental time point. The two other developmental time points each had three samples contributing to the pool. So, three times the quantity of the other individual samples was used to help equalize the time point contribution to the pooled sample. Additionally, 630ng used the entirety of sample #08.

The male RNA pool used 315ng of each sample. This number differs from the 210ng used for the female RNAs so that the two pools would end up with the same total quantity of RNA. However, now that I’ve typed this, this doesn’t matter since the libraries will be equalized before being run on the Illumina HiSeq2500. Oh well. As long as each sample in each pool contributed to the total amount of RNA, then it’s all good.

The two pools were shipped O/N on dry ice.

• Geo_pool_M
• Geo_pool_F

# Sequencing Data – C.gigas Larvae OA

Our sequencing data (Illumina HiSeq2500, 100SE) for this project has completed by Univ. of Oregon Genomics Core Facility (order number 2212).

Samples sequenced/pooled for this run:

 Sample Treatment Barcode 400ppm 400ppm GCCAAT 1000ppm 1000ppm CTTGTA

All code listed below was run on OS X 10.9.5

#!/bin/bash curl -O http://gcf.uoregon.edu:8080/job/download/2212?fileName=lane2_NoIndex_L002_R1_001.fastq.gz curl -O http://gcf.uoregon.edu:8080/job/download/2212?fileName=lane2_NoIndex_L002_R1_002.fastq.gz curl -O http://gcf.uoregon.edu:8080/job/download/2212?fileName=lane2_NoIndex_L002_R1_003.fastq.gz curl -O http://gcf.uoregon.edu:8080/job/download/2212?fileName=lane2_NoIndex_L002_R1_004.fastq.gz curl -O http://gcf.uoregon.edu:8080/job/download/2212?fileName=lane2_NoIndex_L002_R1_005.fastq.gz curl -O http://gcf.uoregon.edu:8080/job/download/2212?fileName=lane2_NoIndex_L002_R1_006.fastq.gz curl -O http://gcf.uoregon.edu:8080/job/download/2212?fileName=lane2_NoIndex_L002_R1_007.fastq.gz curl -O http://gcf.uoregon.edu:8080/job/download/2212?fileName=lane2_NoIndex_L002_R1_008.fastq.gz curl -O http://gcf.uoregon.edu:8080/job/download/2212?fileName=lane2_NoIndex_L002_R1_009.fastq.gz curl -O http://gcf.uoregon.edu:8080/job/download/2212?fileName=lane2_NoIndex_L002_R1_010.fastq.gz curl -O http://gcf.uoregon.edu:8080/job/download/2212?fileName=lane2_NoIndex_L002_R1_011.fastq.gz curl -O http://gcf.uoregon.edu:8080/job/download/2212?fileName=lane2_NoIndex_L002_R1_012.fastq.gz

Renamed all files by removing the beginning of each file name (2112?fileName=) and replacing that with 2212_:

$for file in 2212*lane2_NoIndex_L002_R1_0*; do mv "$file" "${file/#2212?fileName=/2212_}"; done Created a directory readme.md (markdown) file to list & describe directory contents: readme.md $ls *.gz >> readme.md

Note: In order for the readme file to appear in the web directory listing, the file cannot be all upper-case.

Create MD5 checksums for each the files: checkums.md5

$md5 2212* >> checksums.md5 # Sequencing Data – LSU C.virginica MBD BS-Seq Our sequencing data (Illumina HiSeq2500, 100SE) for this project has completed by Univ. of Oregon Genomics Core Facility (order number 2112). Samples sequenced/pooled for this run:  Sample Treatment Barcode HB2 25,000ppm oil ATCACG HB16 25,000ppm oil TTAGGC HB30 25,000ppm oil TGACCA NB3 No oil ACAGTG NB6 No oil GCCAAT NB11 No oil CAGATC All code listed below was run on OS X 10.9.5 Downloaded all 15 fastq.gz files to Owl/web/nightingales/C_virginica: $curl -O http://gcf.uoregon.edu:8080/job/download/2112?fileName=lane1_NoIndex_L001_R1_001.fastq.gz
$curl -O http://gcf.uoregon.edu:8080/job/download/2112?fileName=lane1_NoIndex_L001_R1_002.fastq.gz $curl -O http://gcf.uoregon.edu:8080/job/download/2112?fileName=lane1_NoIndex_L001_R1_003.fastq.gz
$curl -O http://gcf.uoregon.edu:8080/job/download/2112?fileName=lane1_NoIndex_L001_R1_004.fastq.gz $curl -O http://gcf.uoregon.edu:8080/job/download/2112?fileName=lane1_NoIndex_L001_R1_005.fastq.gz
$curl -O http://gcf.uoregon.edu:8080/job/download/2112?fileName=lane1_NoIndex_L001_R1_006.fastq.gz $curl -O http://gcf.uoregon.edu:8080/job/download/2112?fileName=lane1_NoIndex_L001_R1_007.fastq.gz
$curl -O http://gcf.uoregon.edu:8080/job/download/2112?fileName=lane1_NoIndex_L001_R1_008.fastq.gz $curl -O http://gcf.uoregon.edu:8080/job/download/2112?fileName=lane1_NoIndex_L001_R1_009.fastq.gz
$curl -O http://gcf.uoregon.edu:8080/job/download/2112?fileName=lane1_NoIndex_L001_R1_010.fastq.gz $curl -O http://gcf.uoregon.edu:8080/job/download/2112?fileName=lane1_NoIndex_L001_R1_011.fastq.gz
$curl -O http://gcf.uoregon.edu:8080/job/download/2112?fileName=lane1_NoIndex_L001_R1_012.fastq.gz $curl -O http://gcf.uoregon.edu:8080/job/download/2112?fileName=lane1_NoIndex_L001_R1_013.fastq.gz
$curl -O http://gcf.uoregon.edu:8080/job/download/2112?fileName=lane1_NoIndex_L001_R1_014.fastq.gz $curl -O http://gcf.uoregon.edu:8080/job/download/2112?fileName=lane1_NoIndex_L001_R1_015.fastq.gz

Renamed all files by removing the beginning of each file name (2112?fileName=) and replacing that with 2112_:

$for file in 2112*lane1_NoIndex_L001_R1_0*; do mv "$file" "${file/#2112?fileName=/2112_}"; done Created a directory readme.md (markdown) file to list & describe directory contents: readme.md $ls *.gz >> readme.md

Note: In order for the readme file to appear in the web directory listing, the file cannot be all upper-case.

Created MD5 checksums for each fastq.gz file: checksums.md5

\$md5 *.gz >> checksums.md5