Cloning of sugary1 by transposon tagging with Mutator
--Martha G. James and Alan M. Myers

A previous report identified several alleles of sugary1 (su1) in Mutator backgrounds (MNL 66:8). These alleles exhibit a range of phenotypic expression, from wrinkling only at the crown to extremely shrunken and glassy kernels. These variations may be due to background effects, or they may result from position effects of the transposon insertions. Recently, we have identified three additional alleles of su1. These are su1-489 (a gift from Barbara Kloeckener-Gruissem), su1-A1, and su1-A2 (gifts from Mark Alfenito). su1-489 arose in a Mutator background, and su1-A1 and su1-A2 arose in an Ac-background.

Southern hybridization analysis of DNAs from one of the putative Mu-induced alleles, su1-4582, identified a Mu1-homologous, 4.0 kb EcoRI restriction fragment that cosegregated with the mutant phenotype. The 4.0 kb fragment was present in 60 su1-Ref/su1-4582 DNA samples, and absent in 57 su1-Ref/+ sibling DNA samples. A genomic clone corresponding to this 4.0 kb EcoRI fragment was identified and isolated from a bacteriophage lambda library by hybridization with Mu1 (Fig. 1). A hybridization probe from the genomic region flanking the Mu1 insertion ("Su1 probe") also identified a specific 4.0 kb EcoRI fragment in DNAs derived from mutant kernels. Furthermore, the Su1 probe identified similar polymorphisms in DNAs from two independent alleles of su1, su1-7110 and su1-3162. Thus, it is likely that the genomic DNA flanking the Mu1 element in the su1-4582 clone contains at least a portion of the Su1 locus.

A transcript of approximately 3.5 kb was identified in polyadenylated RNA isolated from wild type kernels by hybridization with the Su1 probe. Comparisons with poly A+ RNAs from three of the Mu-induced su1 alleles showed that this transcript was missing in RNA from su1-2412 kernels, was greatly reduced in both size and abundance in su1-4582 kernels, and was slightly larger than the wild type transcript and of roughly equal abundance in RNA from su1-7110 kernels. These differences and their possible relevance to observed phenotypic variations among the transposon-induced alleles are being investigated. A partial cDNA clone homologous to the genomic clone has been isolated from a kernel cDNA library (a gift from Karen Cone).

Figure 1. Restriction map of the cloned genomic EcoRI fragment that cosegregates with su1-4582. The black bar indicates the position of Mu1 within this fragment.

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