09 December 2013
transcriptome and TrinityNext post Previous post
Nice github guide
Finished fourth extraction and pooled all together.
Sent Ap001 DNA sample to Delaware Biotechnology Institute for PacBio and Illumina sequencing.
Sample: Aphaenogaster picea ant collected from Molly Bog, VT
DNA was extracted from 4 sets of 20-40 ants using Qiagen DNeasy kit. Elutions of 300-400 ul per exctraction were pooled for a total volume of about 1300 ul.
Nanodrop quantification showed 15.5 ng/ul with a 260/280 ratio of 1.8. Approximate total of 18 ug DNA.
Took required ethics training course for NSF grant at [https://www.citiprogram.org/]
Writing group from 9-10. Added about 700 word to Methods sections of Aphaenogaster transcriptome paper!
Results of BLASTING known spike-in transcripts to A22 assembly:
|Mean length spike-in (bp)||865.28|
|Mean length assembled (bp)||872.26|
|Mean length missing (bp)||174|
|Mean proportion of original transcript mapped||0.97|
|Mean number assembled transcripts per starting||1.2|
|Proportion bp assembled to starting bp||1.06|
Very nice results!
Set up shell script knit on
home/scripts on workstation, symbolic linked in
usr/local/bin such that globally accessible. Problem that R libraries not installed globally…copied my libraries to
/usr/local/lib/R/site-library/ following Administration and Maintenance of R packages. To globally install package, start R as super-user
and install packages to global library
Confirmed that the
sim.assembly.eval function fails to find any siginificant matches to the VGN Trinity full data assembly that did not include simulated reads!
Using CAP3 or CD-HIT gives similar results (Yang and Smith 2013) so use only CAP3. As I’m now merging contigs within a species, increase similarity threshold to 98%. Previously used 90% when trying to merge orthologs among species.
Found error - removed
trimclip directory with filtered reads, but need these reads for gene expression quantification!
To check reproducibility of script and get these reads, cloned repo into
/home/projects/climate-cascade/projects/ApTranscriptome and ran
to start assembly…
/scripts (master *)$ nohup nice -n 19 knit ApTran_assemble.R &  12512 nohup: ignoring input and appending output to `nohup.out’
Updates to script
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