Molecular analysis of tissue culture-induced white cob mutants in maize (Zea mays.L).

摘要

Plant tissue culture is a mutagenic environment, resulting in a wide array of phenotypic and molecular variation which is observed among regenerated plants and their progenies. There is relative little information available on the molecular basis of tissue culture-induced mutations. In this research, we characterized the molecular basis of five white cob mutants which were identified in progeny of regenerated maize plants. One of the mutations was an allele at the C2 locus, and other mutations were alleles of the P1 locus. The c2 allele resulted from an insertion of 51bp that was found in the second exon of the gene. The inserted sequence consists of a nearly perfect inverted repeat of 43 base pairs and an 8bp duplication of the insertion site. RNA homologous to the c2 gene was not detected in the mutant, likely indicating that the insertion destabilized the transcript. The in silico analysis revealed that the c2 insert had homology to a previously uncharacterized family of transposon-like sequences. The four p1-ww alleles all resulted from epigenetic silencing of this complex locus which consists of six direct repeats. Differential DNA methylation between the mutants and wild-type was found at two sites---an HpaII site at base pair 11167 and a SalI site at base pair 10528. Bisulfite genomic DNA sequencing further identified increased CG and CXG methylation extending across several hundred base pairs surrounding the restriction sites. Asymmetric cytosines were generally unmethylated in these regions. The P transcript was not detected in the mutant, supporting our hypothesis of transcriptional silencing. This research expands our understanding of the molecular basis on tissue culture-induced mutation in plants especially, and more generally, mechanisms by which genomes respond to stress.