title: Polyadenylation state microarray (PASTA) analysis.
abstract: Nearly all eukaryotic mRNAs terminate in a poly(A) tail that serves important roles in mRNA utilization. In the cytoplasm, the poly(A) tail promotes both mRNA stability and translation, and these functions are frequently regulated through changes in tail length. To identify the scope of poly(A) tail length control in a transcriptome, we developed the polyadenylation state microarray (PASTA) method. It involves the purification of mRNA based on poly(A) tail length using thermal elution from poly(U) sepharose, followed by microarray analysis of the resulting fractions. In this chapter we detail our PASTA approach and describe some methods for bulk and mRNA-specific poly(A) tail length measurements of use to monitor the procedure and independently verify the microarray data.
authors.affiliation: Department of Biochemistry and Molecular Biology, Monash University, Clayton, VIC, Australia.
authors.forename: Traude H
authors.initials: TH
authors.lastname: Beilharz
authors.forename: Thomas
authors.initials: T
authors.lastname: Preiss
mesh_terms: D001483:Base Sequence; D018076:DNA, Complementary / Q000096:biosynthesis; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D011072:Poly U / Q000235:genetics; D026723:Polyadenylation; D016133:Polymerase Chain Reaction; D012331:RNA, Fungal / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D012333:RNA, Messenger / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D012441:Saccharomyces cerevisiae / Q000235:genetics
chemical_list: D018076:DNA, Complementary; D012331:RNA, Fungal; D012333:RNA, Messenger; D011072:Poly U; D012321:DNA-Directed RNA Polymerases