Visualization of differential gene expression using a novel method of RNA fingerprinting based on AFLP: analysis of gene expression during potato tuber development.
Using a highly synchronous in vitro tuberization system, in combination with an amplified restriction fragment polymorphism (AFLP)-derived technique for RNA fingerprinting (cDNA-AFLP), transcriptional changes at and around the time point of potato tuberization have been analyzed. The targeted expression analysis of a specific transcript coding for the major potato storage protein, patatin and a second transcript, coding for ADP-glucose pyrophosphorylase, a key gene in the starch biosynthetic pathway is described. This paper confirms that kinetics of expression revealed by cDNA-AFLP analysis are comparable to those found in Northern analysis. Furthermore, this paper reports the isolation and analysis of two tuber-specific transcript-derived fragments (TDFs) coding for the lipoxygenase enzyme, which are differentially induced around the time point of tuber formation. Analysis of the two lox TDFs demonstrates that it is possible to dissect the expression modalities of individual transcripts, not independently detectable by Northern analysis. Finally, it is shown that using cDNA-AFLP, rapid and simple verification of band identity may be achieved. The results indicate that cDNA-AFLP is a broadly applicable technology for identifying developmentally regulated genes.
Bachem
C W
CW
Department of Plant Breeding, Graduate School of Experimental Sciences, Wageningen Agricultural University, The Netherlands. Christian.Bachem/users.pv.wau.nl
van der Hoeven
R S
RS
de Bruijn
S M
SM
Vreugdenhil
D
D
Zabeau
M
M
Visser
R G
RG
D001483:Base Sequence; D002265:Carboxylic Ester Hydrolases; D003001:Cloning, Molecular; D017931:DNA Primers; D018076:DNA, Complementary; D018506:Gene Expression Regulation, Plant; D017343:Genes, Plant; D005821:Genetic Techniques; D051599:Glucose-1-Phosphate Adenylyltransferase; D008969:Molecular Sequence Data; D009713:Nucleotidyltransferases / Q000096:biosynthesis*; D010940:Plant Proteins / Q000096:biosynthesis*; D016133:Polymerase Chain Reaction; D012150:Polymorphism, Restriction Fragment Length; D012333:RNA, Messenger / Q000096:biosynthesis*; D018749:RNA, Plant / Q000032:analysis / Q000096:biosynthesis*; D012689:Sequence Homology, Nucleic Acid; D011198:Solanum tuberosum / Q000235:genetics / Q000254:growth & development / Q000502:physiology*; D013213:Starch / Q000096:biosynthesis; D013997:Time Factors; D014158:Transcription, Genetic
D017931:DNA Primers; D018076:DNA, Complementary; D010940:Plant Proteins; D012333:RNA, Messenger; D018749:RNA, Plant; C043222:patatin protein, Solanum tuberosum; D013213:Starch; D009713:Nucleotidyltransferases; D051599:Glucose-1-Phosphate Adenylyltransferase; D002265:Carboxylic Ester Hydrolases