Western Forest Transcriptome Survey
US FOREST SERVICE

How Do We Count Transcripts?
To measure transcriptome variation, we first isolate RNA from the tissue(s) of interest, then convert the RNA to DNA for sequencing. Multiple samples can be tagged with molecular barcodes to increase sample throughput (as shown in Fig 1).

Figure 1

DNA pools are sequenced by massively parallel sequencing using the Illumina Genome Analyzer (Fig 2).

Figure 2

This technology reads a small portion, or microread, of every DNA in the sample. Millions of microreads are recorded per sample, and these microreads are mapped back to a genome or transcriptome reference to identify which genes are expressed in RNA pool (Fig 3).

Figure 3

Under different experimental conditions, genes may be constantly expressed (as indicated by black microreads that map to two genes), or they may be differentially expressed as shown with red and blue microreads (gene 1 is only expressed in the red treatment). When properly conducted, MPS can be used to discover the sequence of expressed genes and their abundance.

Transcriptomes Evaluated to Date
To date, our collaboration has sequenced a staggering 42 billion bases of DNA from 128 different transcriptome and genome sources. This amount of sequence information dwarfs the human genome, which contains about 3.3 billion bases of DNA.

Transcriptome sequencing by region. PNW, Pacific Northwest Research Station; PSW, Pacific Southwest Research Station; RMRS, Rocky Mountain Research Station.

Transcriptome sequencing by region. PNW, Pacific Northwest Research Station; PSW, Pacific Southwest Research Station; RMRS, Rocky Mountain Research Station.

Data Deposition
Once checked for accuracy, all sequencing data will be deposited to the NCBI Short Read Archive.


Analysis of complete chloroplast genomes (120,000 bp) identifies sites unique to pine species.
Barcoding allows multiple samples to be analyzed simultaneously; note the "CCT" tag Analysis of complete chloroplast genomes (120,000 bp) identifies sites unique to pine species. Field collected transcriptome samples are flash frozen in liquid nitrogen to preserve RNA profiles.
Analysis of complete chloroplast genomes (120,000 bp) identifies sites unique to pine species.

 

Regional Contacts:

Pacific Northwest
Richard Cronn: 541-750-7291,
rcronn@fs.fed.us

Pacific Southwest
Jessica Wright: 530-759-1742,
jessicawright@fs.fed.us

Rocky Mountain
Bryce Richardson: 801-356-5112,
brichardson02@fs.fed.us