NASC's Affymetrix Service

Some sample preparation hints

Here are some things to bear in mind when preparing samples for sending to NASC.

  • When we ran our initial trial experiment, we extracted and prepared our RNA samples using the RNAeasy extraction protocol from Qiagen.
  • We grew our samples under continuous (24 hour) light conditions to reduce circadian influences on the materials. All stocks grown at NASC since 1999 have been bulked under 24-hr light conditions. This also helps to reduce potential maternal circadian effects in seeds grown from NASC stocks.
  • In addition to growth under 24hr light conditions, materials were collected at defined, consistent, and recorded times of day to reduce any material or environmental circadian effects.
  • Material was also collected under defined and recorded light and temperature conditions. Handling was minimal and each individual plant or seedling was snap-frozen immediately upon picking to reduce any stress induction of transcripts upon handling.
  • Plants should be selected at a clearly identified numeric stage as defined by Boyes et al, (2001). Also see our plant ontologies help page. Plants were individually hand picked to align with developmental stagerather than mass harvesting by chronological time. As shown in the reference above, chronological time is a poor determinant of developmental stage for analysis of mutants and comparisons between ecotypes.
  • Light, humidity and a stereotyped watering regime throughout the growing period were followed and recorded (as are compost and other conditions of growth) as described here.
  • Samples were pooled from a minimum of 8 plants to reduce sampling variability.

Affymetrix data of our experiment is published online in our NASCArrays database.

You might want to look at our sample submission form. This will tell you what information we require when you send us samples, and may give some ideas on the factors considered important when designing experiments.

RNA Extraction Protocols

The standard RNA extraction protocols for microarray analysis that most of our customers use are Qiagen and Trizol protocols. Our experience at NASC shows that these protocols are not always ideal for a small number of plant tissues. Below are a list of additional extraction protocols for certain plant tissues;