Insertion mutagenesis and study of transposable elements using a new unstable virescent seedling allele for isolation of haploid petunia lines.
The new unstable virescent seedling (vis) allele of a petunia mutant, that has green leaves but white cotyledons with green revertant spots, was used to identify spontaneously occurring haploid petunia lines with active transposable elements. Endogenous transposons were trapped into the single petunia nitrate reductase structural gene (nia) using chlorate selection on haploid protoplasts. In two mutant lines, the dTph1-like transposable element dTph1-3 was inserted at almost the same position but in opposite orientations in the first exon of the nia gene. In a third mutant, a different transposable element was integrated into the fourth exon. This element, called dTph4, is 787 bp long and has 13 bp terminal inverted repeats of which 12 bp are identical to those of dTph1. Insertion of dTph1-3 and dTph4 results in an 8 bp duplication of the target site, as already described for dTph1. In contrast to dTph1-like elements, dTph4 is present at low copy number in the petunia genome. This can facilitate its use for gene tagging in petunia. The dTph1-3 and dTph4 elements excise frequently, as transposon footprints were found in most of the insertion mutants. The data demonstrate that haploid petunia is an excellent system for gene tagging and for the study of transposable elements.
Renckens
S
S
Laboratorium Genetische Virologie, Vrije Universiteit Brussel, Sint-Genesius-Rode, Belgium.
De Greve
H
H
Beltrán-Herrera
J
J
Toong
L T
LT
Deboeck
F
F
De Rycke
R
R
Van Montagu
M
M
Hernalsteens
J P
JP
D000483:Alleles; D001483:Base Sequence; D002736:Chloroplasts / Q000648:ultrastructure; D003001:Cloning, Molecular; D004251:DNA Transposable Elements / Q000235:genetics*; D018744:DNA, Plant / Q000032:analysis; D005091:Exons / Q000235:genetics; D017343:Genes, Plant / Q000235:genetics; D006238:Haploidy; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional / Q000379:methods*; D050901:Nitrate Reductase; D009565:Nitrate Reductases / Q000235:genetics; D010641:Phenotype; D010944:Plants / Q000201:enzymology / Q000235:genetics*; D011523:Protoplasts; D012091:Repetitive Sequences, Nucleic Acid / Q000235:genetics; D017422:Sequence Analysis, DNA
D004251:DNA Transposable Elements; D018744:DNA, Plant; D009565:Nitrate Reductases; D050901:Nitrate Reductase