Single-cell laser-capture microdissection and RNA amplification.
Generating gene-expression profiles from laser-captured cells requires the successful combination of laser-capture microdissection, RNA extraction, RNA amplification, and microarray analysis. To permit single-cell gene-expression profiling, the RNA amplification method has to be sufficiently powerful to bridge the gap between the amount of RNA available from a single cell to what is required by the microarray, a gap that spans 5 to 6 orders of magnitude. This chapter focuses on the amplification of RNA using a two-round T7 RNA amplification method. The protocols described are adapted for laser-captured material and have been used to generate gene expression profiles from single laser-captured cells.
Kamme
Fredrik
F
Pharmaceutical Research & Development, Johnson & Johnson, San Diego, CA, USA.
Zhu
Jessica
J
Luo
Lin
L
Yu
Jingxue
J
Tran
Da-Thao
DT
Meurers
Bernhard
B
Bittner
Anton
A
Westlund
Karin
K
Carlton
Susan
S
Wan
Jackson
J
D004926:Escherichia coli / Q000235:genetics; D020869:Gene Expression Profiling / Q000379:methods; D007834:Lasers; D042282:Microdissection / Q000295:instrumentation / Q000379:methods*; D021141:Nucleic Acid Amplification Techniques / Q000295:instrumentation / Q000379:methods*; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods; D012313:RNA / Q000235:genetics*
D012313:RNA