IDENTIFICATION OF A REGION OF 5' NONCODING SEQUENCE REQUIRED FOR MAXIMAL TRANSCRIPTION OF A ZEIN GENE
Abstract
In order to delimit sequences that are required for the transcription of a zein gene that encodes a Mr 19,000 zein, a progressive series of deletions in the 5' noncoding sequence of a genomic zein clone, pgZ19abl, were generated. The transcriptional activity of the 5' deletion series was analyzed in two heterologous plant systems. The Ti plasmid was used to introduce the modified zein genes into the sunflower genome. Electroportation was used to facilitate the uptake of plasmid DNA by carrot protoplasts; the plasmids contained the truncated promoter regions from the 5' deletion series linked to the protein coding sequences of the bacterial chloramphenicol acetyl transferase (CAT) gene. Transcriptional activity was analyzed by S1 nuclease mapping of zein specific tumor transcripts and CAT enzyme activity assays of carrot protoplast extracts, respectively. These analyses led to identification of a region of upstream sequence that is required for maximal transcription of the zein gene. This region is delimited by positions -337 and -125 with respect to the mRNA cap site. Very low, but detectable levels of transcription were directed by zein genes with only 125 or 79 pb of upstream sequence, suggesting that regulatory regions 5' to the CAAT and TATA boxes exert a quantitative effect on transcription. Examination of the sequence in this regulatory region showed five regions that shared imperfect homology with the SV40 enhancer "core"-like sequence. Additionally, alternating purine/pyrimidine repeats and ordered purine/pyrimidine repeat units were identified in this region.
Degree
Ph.D.
Subject Area
Botany
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