Identification of differentially expressed genes in anagen dermal papilla by suppression subtractive hybridization.
BACKGROUND
We constructed a cDNA subtractive library of dermal papilla cells (DPCs) in anagen with suppression subtractive hybridization (SSH) technique and clone differentially expressed genes related to DPCs in anagen.


METHODS
Total mRNA was isolated from DPCs of anagen and telogen follicles. Moreover, single-strand (ss) and double-strand (ds) cDNAs were synthesized in turn using SMART PCR cDNA synthesis technology. ds cDNAs then were digested with Rsa I and divided into two groups, and ligated to the specific adaptor 1 and adaptor 2R, respectively. After cDNAs were hybridized with each other twice and underwent two rounds of nested PCR. PCR products were ligated with arms of T/A plasmid vectors to set up the subtractive library. Selected clones were demonstrated by reverse Northern blot and sequenced. The acquired sequence data were aligned against the Genbank nucleotide database.


RESULTS
cDNA subtractive library of DPCs in anagen follicles was set up successfully with high subtractive efficiency. Thirty-five genes were identified in this study with 22 known functional genes and 13 unknown functional genes.


CONCLUSIONS
All results confirm the effectiveness and sensitivity of SSH in detecting differentially expressed genes from a small amount of clinical samples. Information about such alterations in gene expression could be useful for elucidating the genetic events in hair follicle growth regulation.
Yang
Xi-chuan
XC
Department of Dermatology, Southwest Hospital of Third Military Medical University, Chongqing 400038, China.
Hao
Fei
F
Song
Zhi-qiang
ZQ
Cheng
Bo
B
Yang
Wei-bing
WB
Zhong
Bai-yu
BY
Xiang
Ming-ming
MM
D000328:Adult; D000506:Alopecia Areata / Q000235:genetics*; D018076:DNA, Complementary / Q000032:analysis*; D005260:Female; D015723:Gene Library; D006197:Hair; D018859:Hair Follicle / Q000737:chemistry*; D006801:Humans; D009693:Nucleic Acid Hybridization; D016133:Polymerase Chain Reaction
D018076:DNA, Complementary