Community Research and Development Information Service - CORDIS

Primers and protocols for PCR amplification of selected genes whose expression is indicative for dormancy or hardiness in pine

We selected 9 pine genes that are likely to be involved in pine cold hardiness development and developed RT-PCR primers to allow rapid and robust analysis of the quantitative expression levels of these genes. It concerns
- dehydrin Psdhn1
- dehydrin Psdhn2
- dehydrin family Psdhn3-8 (group-specific)
- dehydrin Psdhn5
- dehydrin Psdhn3/7 (double detection)
- a stress inducible protein
- a Tubulin
- Glucose-6-Phosphate Dehydrogenase
- Ribosomal 18S RNA.
We developed a reliable protocol that takes into account the difficulties of the tissue type (dormant buds). The protocol starts with a CTAB RNA isolation. Preparations were DNAseI and used for cDNA synthesis. Dilutions of this cDNA were used for Realtime PCR. Product formation was measured using the iCycler system (BIORAD Laboratories, The Netherlands). Relative changes in expression were calculated using the Gene Expression Macro (Version 1.1) supplied by BIORAD.

These primers can be used for quantitative analysis of stress related gene expression.
The can be used in scientific research.
In addition, they may serve as content for a molecular diagnostic assay for hardiness of pine seedlings

Contact

Monique VAN WORDRAGEN, (Programme Co-ordinator)
Tel.: +31-31-7475114
Fax: +31-31-7475347
E-mail
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