RNA sequencing (Sciuridae)

Knowledge graph centered on RNA sequencing (Sciuridae) with 61 nodes and 135 connections. Top connected: Rocky Mountain Biological Laboratory, Gothic, Marmota, Yellow-bellied Marmot, Crested Butte.

Method synopsis

RNA extraction from blood samples, library preparation, and high-throughput sequencing on Illumina platforms followed by read mapping and differential gene expression analysis using mixed models.

Synthesized from method descriptions across 2 papers using this protocol.

Procedure from a recent peer-reviewed implementation

Steps below were extracted from the most recent peer-reviewed implementation of this protocol in the corpus — Gene expression shifts in yellow-bellied marmots prior to natal dispersal (2019), Behavioral Ecology. Implementations in other papers (listed below) may differ.

1. Collect blood samples
   During each yearling capture, collected 1 mL whole blood and preserved it in 2.5 mL PAXgene Blood RNA solution
   Quantities: 1 mL whole blood per individual, preserved in 2.5 mL PAXgene solutionDuration: During capture eventsConditions: At time of live capture
   Equipment: PAXgene Blood RNA solution
2. Extract RNA from blood samples
   Samples were incubated, frozen, and extracted according to PAXgene Blood RNA kit manufacturers' instructions. Only samples taken during the dispersal period up to 10 days after the colony's pup emergence were included in subsequent steps.
   Quantities: 51 samples total analyzedDuration: Following manufacturer protocolsConditions: Laboratory conditions following PAXgene protocols
   Equipment: PAXgene Blood RNA kit
3. Remove globin transcripts and assess RNA quality
   Removed globin transcripts using the GLOBINclear kit for mouse/rat (Ambion, Austin, MA) and RNA quality was assessed with an Agilent 2100 Bioanalyzer
   Quantities: All 51 RNA samples processedDuration: Following manufacturer protocolsConditions: Laboratory conditions

Typical Equipment

• TruSeq Library Prep Kit v2
• Illumina HiSeq2000
• Illumina HiSeq4000
• TopHat2
• HT-Seq

Output Measurements

• gene expression levels
• transcript counts
• differential expression statistics

Papers Using This Protocol (2)

Species Studied With This Protocol

Places Where Used