cDNA Cloning and Gene Characterization of Large and Small Subunits of Ribonucleotide Reductase in Soybean
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cDNA Cloning and Gene Characterization of Large and Small Subunits of Ribonucleotide Reductase in Soybean by Xinsheng Xiong Joe R. Cowles, Committee Chairman Department of Biology Abstract Ribonucleotide reductase (RNR) reduces four ribonucleoside diphosphates to corresponding deoxyribonucleoside diphosphates, which are transformed into deoxyribonucleoside triphosphates, substrates for DNA polymerase. By controlling the supply and balance of deoxyribonucleoside diphosphates, RNR regulates DNA synthesis. RNR in E. coli and in animals consists of two identical large and two identical small subunits. Until recently, little was known about RNR in plants. For cloning RNR cDNA in plants, soybean (Glycine max) cDNAs were amplified with highly degenerate primers and the Rapid Amplification of cDNA Ends techniques. The cDNAs encoding two complete large subunits, one partial large subunit and one complete small subunit of RNR in soybean were cloned and sequenced. The RNR large subunits in soybean contain a motif with 20 amino acids, which appears to be specific for the RNR large subunits in plants. Southern hybridization results imply that a gene family encodes at least three different large subunits of RNR in soybean, and that a single gene encodes the small subunit. The presence of three different large subunits of RNR in soybean suggests that RNR complex in some plants may have a non-homodimer structure; alternatively, some plants may have different RNR isozymes. Northern hybridization results show that RNR large and small subunit genes in soybean are expressed both in dark-grown and light-grown seedlings, and that light does not increase RNR mRNA levels. Multiple poly(A) sites and different lengths of the 3â untranslated regions were found in cDNAs encoding some subunits of RNR in soybean. The same cis-acting elements may imprecisely locate some multiple poly(A) sites in plants.
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