The DNAs of the plasmids pTac39 and pZmBIB, which contain the wheat chloroplast DNA BunzHI fragment 2 and the maize chloroplast DNA BumHI fragment 9 respectively, were analysed by electron microscopy and by Northern blotting. Both plasmids contain the gene for the large subunit of ribulose bisphosphate carboxylase (LS gene). Comparison of thc DNAs of the two species by partial denaturation and heteroduplex analysis shows extended homologies interspersed with a fine pattern of non-homology. On analysis of wheat-RNA . maize-DNA hybrids a region of non-homology at the 3' end of the LS mRNA of about 200 bases was found. Analysis of wheat D N A . RNA hybrids showed that the wheat LS mRNA contains 1700 1 50 bascs and another hybrid, containing 2400 bases was also shown to occur in a position adjacent to that formed by the LS mRNA. Binding of Escherichiu coli RNA polymerase and the start of transcription in vicro occurred at welldefined sites, which are located between or close to the positions where mRNA . DNA hybrids are found. A modified technique for the preparation and fractionation by electrophoresis of glyoxdlated RNA is described.The structural gene for the large subunit of the enzyme ribulose bisphosphate carboxylase (LS gene) has been localised in chloroplast DNA (ctDNA) using cloned material from maize [I,21, Clzlumydomonas [3], spinach [4] and wheat ( [ 5 ] and A. Doherty, C. Bowman, J. Gray, and T. A. Dyer, unpublished results). Furthermore the sequences of the maize [7] and spinach [8 J LS genes and also the presence of a second gene with a size of about 2200 bases close to the LS gene have been described [9]. The sequence analyses 'indicated the start of mRNA transcription and the length of the protein-coding sequence, but with maize the promoter sequences and those which cause the termination of transcription were not clearly identified. We describe and compare here the organization of wheat and maize ctDNA in the region containing the LS gene. Electron micrographs of hybrids between the wheat mRNA and these DNAs were used in the determination of the precise position and size of the major transcripts. The results so obtained were correlated with those obtained from Northern blots. In addition the positions of probable promoter sequences were determined by observing the binding and transcription in vitro of the wheat and maize DNAs by Escherichia c d i RNA polymerase. The results make it possible to align the gene and control sequences in the region of the LS gene in these two types of ctDNA.
MATERIALS A N D METHODS
Plasmid DNAsThe BamHI fragment 2 of wheat ctDNA, which contains the LS gene, was cloned in pBR322 to give pTac39 (Doherty, Bowman, Gray, and Dyer, unpublished results). The corresponding maize fragment (5amHI 9) was also cloned in pBR322 giving pZmBlB [7]. In some experiments use was made of a BglII subfragment excised from the maize BumHI fragment 9 [2] and in other experiments an EcoRI subfrdgment excised from the wheat BarnHI fragment 2, cloned in pBR322, giving pTac509 was used. Superco...