Addition-deletion mutations in transgenic Arabidopsis thaliana generated by the seed co-cultivation method

We have characterized three mutant alleles of the CER2 gene of Arabidopsis thaliana by sequencing polymerase chain reaction products of this gene generated with template DNA isolated from lines MK1, BRL7, and BRL17. Sequence analysis of the amplification product from line BRL17 revealed a 17-bp deletion in the CER2 gene. The CER2 gene of BRL7 differs from the wild-type sequence by a 2-base substitution and 2-base insertion. As both of these lines were isolated from the transformant populations generated by Dr. K. Feldmann and his collaborators, we suggest that these small rearrangements may be caused by unsuccessful T-DNA insertions. Comparative sequence analysis of the sequence of line MK1 and the wild type revealed a single nucleotide substitution located 360 bp downstream from the intron-exon junction that changes a tryptophan triplet TGG to TGA; i.e., an opal non-sense mutation. In accordance with these observations, the cer phenotype of line MK1 was complemented by transformation with a fusion construct of the CaMV 35S promoter and the CER2 structural gene. Comparative analysis of the deduced amino acid sequences encoded by the CER2 gene from Arabidopsis and by the glossy2 gene from Zea mays revealed a significant similarity between them. This suggests that these gene products may have a similar biochemical function.