Generation and use of a tailored gene array to investigate vascular biology.
Vasculogenesis, angiogenesis and vascular remodelling are complex processes where the fate of several cell types is determined by different signalling networks. Many of these networks ultimately function by changing the abundance of RNA transcripts within the cells which constitute blood vessel walls. Researchers can now map these transcript abundance changes using gene array technology. In this review, we describe the design, production and use of a gene array specifically tailored to investigate vascular biology. We describe the advantages of tailored gene arrays, and give detailed protocols based on our experience to allow the reader to use such gene arrays to generate meaningful data. We list the issues to consider when choosing and verifying the genes and splice variants included in an array, and describe our use of Arabidopsis sp. RNA spikes for quality control. We present data that illustrates the absolute necessity for both technical and biological replicates to be incorporated in the design of gene array experiments using primary cells such as HUVECS. Finally, we describe methods for the normalisation and interpretation of the data that gene arrays produce. The approach to gene array technology described here is easily within reach of the budget and expertise of most academic research groups.
Evans
Amanda L
AL
Reproductive Molecular Research Group, University of Cambridge, Tennis Court Rd, Cambridge, UK.
Sharkey
Andrew S
AS
Saidi
Samir A
SA
Print
Cristin G
CG
Catalano
Roberto D
RD
Smith
Stephen K
SK
Charnock-Jones
D Stephen
DS
D002320:Cardiovascular Physiological Phenomena; D005786:Gene Expression Regulation / Q000502:physiology*; D018919:Neovascularization, Physiologic / Q000235:genetics / Q000502:physiology*; D020411:Oligonucleotide Array Sequence Analysis; D016133:Polymerase Chain Reaction; D015398:Signal Transduction / Q000502:physiology