Regulation of anthocyanin biosynthetic genes introduced into intact maize tissues by microprojectiles

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Abstract
We have employed microprojectiles to deliver genes involved in anthocyanin biosynthesis to cells within intact aleurone and embryo tissues of maize. Clones of the A1 or Bz1 genes were introduced into aleurone tissue that lacked anthocyanins due to mutations of the endogenous A1 or Bz1 gene. Following bombardment, cells within the aleurone developed purple pigmentation, indicating that the mutation in the a1 or bz1 genotypes was corrected by the introduced gene. To analyze the expression of these genes in different genetic backgrounds, chimeric genes containing the 5′ and 3′ regions of the A1 or Bz1 genes fused to a luciferase coding region were constructed. These constructs were introduced into aleurones of genotypes carrying either dominant or recessive alleles of the C1 and R genes, which are known to regulate anthocyanin production. Levels of luciferase activity in permissive backgrounds ( C1, R ) were 30- to 200-fold greater than those detected in tissue carrying one or both of the recessive alleles ( c1, r ) of these genes. These results show that genes delivered to intact tissues by microprojectiles are regulated in a manner similar to the endogenous genes. The transfer of genes directly to intact tissues provides a rapid means for analyzing the genetic and tissue-specific regulation of gene expression.