Using quantitative PCR to identify kinesin-3 genes that are upregulated during growth arrest in mouse NIH3T3 cells.
Most cells in our body form a single primary cilium when entering growth arrest. During the past decade, a number of studies have revealed a key role for primary cilia in coordinating a variety of signaling pathways that control important cellular and developmental processes. Consequently, significant effort has been directed toward the identification of genes involved in ciliary assembly and function. Many candidate ciliary genes and proteins have been identified using large-scale "omics" approaches, including proteomics, transcriptomics, and comparative genomics. Although such large-scale approaches can be extremely informative, additional validation of candidate ciliary genes using alternative "small-scale" approaches is often necessary. Here we describe a quantitative PCR-based method that can be used to screen groups of genes for those that are upregulated during growth arrest in cultured mouse NIH3T3 cells and those that might have cilia-related functions. We employed this method to specifically search for mouse kinesin-3 genes that are upregulated during growth arrest and identified three such genes (Kif13A, Kif13B, and Kif16A). In principle, however, the method can be extended to identify other genes or gene families that are upregulated during growth arrest.
Thorsteinsson
Rikke I
RI
Department of Biology, Section of Cell and Developmental Biology, University of Copenhagen, Universitetsparken 13, DK-2100 Copenhagen OE, Denmark.
Christensen
Søren T
ST
Pedersen
Lotte B
LB
D000818:Animals; D018929:Cell Culture Techniques; D005786:Gene Expression Regulation; D016547:Kinesins / Q000235:genetics* / Q000378:metabolism; D051379:Mice; D041681:NIH 3T3 Cells / Q000502:physiology; D016133:Polymerase Chain Reaction / Q000295:instrumentation / Q000379:methods*; D020033:Protein Isoforms / Q000235:genetics* / Q000378:metabolism; D015854:Up-Regulation
D020033:Protein Isoforms; D016547:Kinesins