A plasmid borne larvicidal crystal protein gene from B.thuringiensis subspecieskurstaki was cloned inEscherichia coli using a specific 20-mer oligonucleotide probe. The gene expressed inE. coli at a high level. TransgenicE. coli cells produced large irregular bodies which looked bright under phase contrast microscopy. The phase bright bodies released by sonic disruption of cells could be pelleted by centrifugation. Toxicity trials on the larvae ofSpodoptera litura showed that the pellet was antifeedant and toxic to the larvae. The supernatant was only mildly antifeedant. Even short term feeding of larvae on the toxin delayed the onset of pupation.

Hierarchical clustering and similarity coefficients of pairwise alignments of the published nucleotide sequences of 27nifH genes suggest thatnif genes are as ancient as the archaebacteria and clostridia. The positions ofnifHl ofMethanococcus thermolithotrophicus, nifH3 ofClostridium pasteurianum, nifH3 ofAzotobacter vinelandii andnifH ofFrankia suggest that a variety of lateral transfers may have occurred during evolution ofnifH gene. The genes for type 3 nitrogenase ofA. vinelandii may have diverged early from methanogens and clostridia. A high similarity coefficient with the derived amino acid sequence of type 3 nitrogenase suggests the presence of a functionally similar enzyme inC. pasteurianum. The type 2 nitrogenase genenifH2 of azotobacters seems to have originated recently from the genenifHl for conventional type I nitrogenase. RhizobialnifH genes comprise two closely related but discrete clusters that are in consonance with the plasmid or chromosomal location ofnif genes. The chromosomal and plasmid locatednifH of rhizobia seem to have evolved independently but contemporaneously.

A 5-5-kbpHindIII fragment carryingpsbE-F-L-J operon from the chloroplast DNA ofPopulus deltoides was cloned and nucleotide sequence was determined for a 1672-bp region. The coding regions showed more than 90% homology at nucleotide sequence level with plastid-encodedpsbE, psbF, psbL andpsbJ genes of higher plants. Pairwise alignments ofpsbE, psbF, psbL andpsbJ coding regions of poplar with published sequences from other plants were carried out to analyse the nature of nucleotide substitutions. The 5’ and 3’ untranslated regions of the genes revealed high variability among different organisms both in terms of homology and the number of nucleotides. Northern hybridization data indicated that all four genes ofpsbE-F-L-J operon were transcribed as a single tetracistronic message which was not subjected to further processing into smaller transcripts. The transcript showed quantitative increase in mature leaves.

We report the cloning and nucleotide sequence analysis ofpsbD/C operon from a dicotyledonous tree species,Populus deltoides (poplar). The coding regions ofpsbD andpsbC and deduced amino acid sequences show very high homology with those from other higher plants. In pairwise alignment of the gene sequences,P. deltoides clustered with dicotyledonous annuals rather than withPinus, the only other tree whosepsbD/C nucleotide sequence is available. Comparison of several reported sequences showed that synonymous substitutions were distributed in bothpsbD andpsbC uniformly, throughout the length of the genes. The frequency of nonsynonymous substitutions located in the amino-terminal endof psbD was distinctly higher, suggesting a lower degree of structural constraints in this region of the encoded D2 protein. The arrangement of reading frames and Northern analysis suggest that organization and expression ofpsbD/C operon inP. deltoides is similar to that in other higher plants.

Genes that code for proteins expressed at high and low levels in plants were classified into separate data sets. The two data sets were analysed to identify the conserved nucleotide sequences that may characterize genes with contrasting levels of expression. The AUG context that characterized the highly expressed genes is (A/C)N₂AAN₃(A/T)T(A/C) AACAATGGCTNCC(T/A)CNA(C/T)(A/C). The data set of highly expressed genes shows overrepresentation of codons for alanine at the second position and serine at the third and fourth positions after the translation initiation codon. The characteristic transcription initiation site in the highly expressed genes is CAN(A/C)(A/C)(C/A)C(C/A)N₂A(C/A). The promoter region is characterized by two tandemly repeated TATA elements, sometimes with one and rarely with two point mutations in the highly expressed genes. Besides the two tandemly repeated TATA elements, the promoter context in the highly expressed genes is overrepresented by C, C and G at the -3, -1 and+9 positions respectively. The characteristic TATA motif in the highly expressed plant genes is (T/C)(T/A)N₂TCACTATATATAG. Most of these features are not present in the genes ubiquitously expressed at low levels in plants.