Resistance to streptomycin (Sm) of Euglena gracilis chloroplasts can be due to a single C to T transition of the 16S rRNA gene in an invariant position which is equivalent to C912 of the Escherichia coli 16S rRNA. Since Euglena chloroplasts cannot be transformed we introduced, by site‐directed mutagenesis, a C912 to T transition in the cloned rrnB operon (pKK3535) of E. coli and used this new construct (pEM109) in transformation experiments. Transformed E. coli cells were selected for Sm resistance by colony plating and stepwise increase of Sm up to 25 micrograms/ml of culture medium. Several Sm‐resistant colonies were obtained. Ribosomes were isolated from pEM109‐transformed Sm‐resistant and pKK3535‐transformed Sm‐sensitive cells. The ribosomes were assayed in vitro for Sm‐induced misreading of poly(U) mRNA. We isolated 16S rRNA and sequenced the crucial RNA region by reverse transcription. The results clearly show that ribosomes from Sm‐resistant cells correctly read the poly(U) mRNA in the presence of 25 micrograms Sm/ml of reaction mixture and the 16S rRNA contains the C912 to U transition. We conclude that C912 is involved in a translation step(s) which is (are) sensitive to streptomycin.

We characterize a 1.95 kb transcription product of the Euglena gracilis chloroplast DNA fragment Eco-N + Q by S1 nuclease analysis and DNA sequencing and show that it is the product of three splicing events. Exon 1 (0.

We characterize a DNA segment of the Euglena gracilis chloroplast DNA fragment Eco . N by nucleotide sequencing and S1 nuclease analysis. We show that this region, which is upstream of the previously sequenced tuf A gene, contains the genes for the ribosomal proteins S12 and S7. The gene arrangement is 5'-rps 12-80 bp spacer-rps 7-174 bp spacer-tuf A, somewhat similar to the str operon of E. coli. The chloroplast S12 and S7 proteins contain 124 and 155 aminoacids, respectively, and are to 68% and 38% homologous with the corresponding E. coli proteins. The region is transcribed into a distronic mRNA of about 1.1 to 1.2 kb. The rps 12 and rps 7 genes, contrary to the tuf A gene, are not split.

We sequenced the chloroplast 16S rRNA gene of two Euglena gracilis mutants which contain streptomycin-resistant chloroplasts (Smr 139.12/4 and Smr 139.20/2). These mutants are known to contain a single intact rrn operon per circular chloroplast genome. Nucleotide sequence comparison between a 16S rRNA gene of wild type Euglena gracilis, strain Z, with streptomycin-sensitive chloroplasts, and the 16S rRNA gene of both Smr-strains reveals a single base change (C to T) at position 876. This position is equivalent to the invariant position 912 of the E. coli 16S rRNA gene. The analogous position is also conserved in all chloroplast small subunit RNA genes from lower and higher plants sequenced so far. Light dependent protein synthesis with purified chloroplasts from streptomycin-resistant cells is not inhibited by streptomycin. Based on the results reported here we postulate linkage between the observed point mutation on the 16S rRNA gene and streptomycin-resistance of chloroplast 70S ribosomes.

A cDNA and a genomic DNA library from soybean (Glycine max L.) were used to identify and sequence two genes coding for the alpha-subunit of the translation elongation factor eEF-1. Within the coding part, the two genes (tefS1 and tefS2) diverge in 80 wobble positions thus yielding an identical protein composed of 447 amino acids. The soybean protein has about 95% similarity with eEF-1 alpha proteins of Arabidopsis thaliana and tomato. Both genes S1 and S2 contain, within the coding part at a site seemingly unique to higher plants, a single short intron of 86 and 116 nucleotides, respectively. The untranslated leader part of both genes is interrupted by a large intron (partially sequenced). Genes S1 and S2 are transcribed in young leaves. cDNA and gene-specific oligonucleotide probes interact with a unique transcript of close to 1.9 kb. Northern hybridization studies using RNAs from dark- and light-grown seedlings show that light sharply increases the level of stable transcripts (1.9 kb). A peak value is measured after about 3 h of illumination, afterwards the transcript concentration drops to about 10% of the peak value. Genes S1 and S2 follow a similar transcription pattern in developing seedling leaves, which is distinct from that of the rbcS genes measured in parallel experiments. According to northern results, S1 transcripts are more abundant in leaves at all measured stages of development than S2 transcripts.