title: A novel sample labeling method with restriction display PCR for 60-mer oligonucleotide microarray.
abstract: OBJECTIVE
  To investigate the value of restriction display PCR (RD-PCR) as a novel and expedient sample labeling method for high-density 60-mer oligonucleotide microarray.
  
  
  METHODS
  Peripheral blood samples from three volunteers were collected and the total RNA was extracted from the peripheral blood mononuclear cells and labeled with RD-PCR protocol, followed by hybridization with Agilent Human 1B oligonucleotide microarrays in a two-color comparison format. The RNA from the same subject was divided into two aliquot and labeled with Cy3 and Cy5 respectively. The spots with significant difference between the foreground and local background intensities and those without significant difference between Cy5 and Cy3 signal intensities were selected for analysis. SPSS software was used to perform the statistical tests and plot generation. VSN packages were used under R language to remove the systematic array and dye biases.
  
  
  RESULTS
  Totally 8744 common spots of the 3 microarrays were evaluated. The results demonstrated that RD-PCR could be a promising novel method for efficient labeling of microarray samples. Further analysis indicated the presence of adjustable biases derived from the array and incorporated dye in the labeling processes. The RD-PCR labeling showed better performance than the conventional approaches in regards to reproducibility of the quantitative signals for gene intensity and capability to label RNAs of lowly expressed genes.
  
  
  CONCLUSIONS
  Given the evidence of the feasibility of using RD-PCR labeling in the field of high-density long oligonucleotide microarray, further optimization of the protocol may unleash the full potential of this novel labeling method.
authors.affiliation: Institute of Genetic Engineering, Southern Medical University, Guangzhou 510515, China. moxiaoyang@yahoo.com.cn
authors.forename: Xiao-yang
authors.initials: XY
authors.lastname: Mo
authors.forename: Wen-li
authors.initials: WL
authors.lastname: Ma
authors.forename: Ling
authors.initials: L
authors.lastname: Li
authors.forename: Rong
authors.initials: R
authors.lastname: Shi
authors.forename: Bao
authors.initials: B
authors.lastname: Zhang
authors.forename: Qiu-lin
authors.initials: QL
authors.lastname: Xu
authors.forename: Hai-yan
authors.initials: HY
authors.lastname: Zhang
authors.forename: Wen-ling
authors.initials: WL
authors.lastname: Zheng
mesh_terms: D006801:Humans; D007963:Leukocytes, Mononuclear / Q000378:metabolism*; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D015345:Oligonucleotide Probes / Q000737:chemistry; D016133:Polymerase Chain Reaction / Q000379:methods*; D013194:Staining and Labeling
chemical_list: D015345:Oligonucleotide Probes