Molecular organization of great millet (Sorghum vulgare) DNA

Approximately 52% of the nuclear genome of great millet(Sorghum vulgare) consists of repetitive DNA which can be grouped into very fast, fast and slow components. The reiteration frequencies of the fast and slow reassociating components are {dy7000} and 92 respectively. Approximately 90% of the genome consists of repeated sequences interspersed amongst themselves and with single copy sequences. The interspersed repeat sequences are of three sizesviz. > 1·5 kilobase pairs, 0·5–1·0 kilobase pairs and 0·15–0·30 kilobase pairs while the size of the single copy sequences is 3·0 kilobase pairs. Hence the genome organization of great millet is essentially of a mixed type NCL communication No. 3527.     

Additional evidence implicating Triticum searsii as the B-genome donor to wheat

In vitro DNA:DNA hybridizations and hydroxyapatite thermal-elution chromatography were employed to identify the diploid wheat species ancestral to the B genome of Triticum turgidum. ³H-T. turgidum DNA was hybridized to the unlabeled DNAs of T. urartu, T. speltoides, T. sharonensis, T. bicorne, T. longissimum, and T. searsii. ³H-Labeled DNAs of T. monococcum and a synthetic tetraploid AADD were hybridized with unlabeled DNAs of T. urartu and T. searsii to determine the relationship of the A genome of polyploid wheat and T. urartu. The heteroduplex thermal stabilities indicated that T. searsii was most closely related to the B genome of T. turgidum (AB) and that the genome of T. urartu and the A genome have a great deal of base-sequence homology. Thus, it appears that T. searsii is the B-genome donor to polyploid wheat or a major chromosome donor if the B genome is polyphyletic in origin.Published with the approval of the Director of The West Virginia Agricultural Experiment Station as Scientific Paper No. 1837.     

The ribosomal RNA genes from chloroplasts of mustard (Sinapis alba L.): mapping and sequencing of the leader region

Summary The genes coding for rRNAs from mustard chloroplasts were mapped within the inverted repeat regions of intact ctDNA and on ctDNA fragments cloned in pBR322. R-loop analysis and restriction endonuclease mapping show that the genes for 16S rRNA map at distances of 17 kb from the junctions of the repeat regions with the large unique region. The genes for 23S rRNA are located at distances of 2.8 kb from the junctions with the small unique region. Genes for 4.5S and 5S rRNA are located in close proximity to the 23S rRNA genes towards the small unique region. DNA sequencing of portions of the 5 terminal third from the mustard 16S rRNA gene shows 96–99% homology with the corresponding regions of the maize, tobacco and spinach chloroplast genes. Sequencing of the region proximal to the 16S rRNA gene reveals the presence of a tRNA^Val gene in nearly the same position and with identical sequence as in maize, tobacco and spinach. Somewhat less but still strong homology is also observed for the tDNA ^Val/16S rDNA intercistronic regions and for the regions upstream of the tRNA^Val gene. However, due to many small and also a few larger deletions and insertions in the leader region, common reading frames coding for homologous peptides larger than 44 amino acids can not be detected; it is therefore unlikely that this region contains a protein coding gene.     

Homology between the ribosomal DNA of Escherichia coli and mitochondrial DNA preparations of maize is principally to sequences other than mitochondrial rRNA genes

E. coli ribosomal DNA has been used to probe maize mitochondrial DNA. It hybridizes primarily with chloroplast ribosomal DNA sequences and with fungal and bacterial sequences which may contaminate the mtDNA preparations. It also hybridizes to the chloroplast 16S ribosomal RNA gene sequence present in the mitochondrial genome (1) as well as to the mitochondrial 18S ribosomal RNA gene sequence. Weak sequence homology was detected between E. coli rDNA and the mitochondrial 26S ribosomal RNA gene.     

Assessment of a model for intron RNA secondary structure relevant to RNA self-splicing--a review

A widespread class of introns is characterized by a particular RNA secondary structure, based upon four conserved nucleotide sequences. Among such "class I" introns are found the majority of introns in fungal mitochondrial genes and the self-splicing intron of the large ribosomal RNA of several species of Tetrahymena. A model of the RNA secondary structure, which must underlie the self-splicing activity, is here evaluated in the light of data on 16 further introns. The main body or "core structure" of the intron always consists of the base-paired regions P3 to P9 with the associated single-stranded loops, with P2 present also in most cases. Two minority sub-classes of core structure occur, one of which is typical of introns in fungal ribosomal RNA. Introns in which the core structure is close to the 5' splice site all have an internal guide sequence (IGS) which can pair with exon sequences adjacent to the 5' and 3' splice sites to align them precisely, as proposed by Davies et al. [Nature 300 (1982) 719-724]. In these cases, the internal guide model allows us to predict correctly the exact location of splice sites. All other introns probably use other mechanisms of alignment. This analysis provides strong support for the RNA splicing model which we have developed.     

Structural properties of phospholipids in the rat liver inner mitochondrial membrane. A P-NMR study

1. 1. The ³¹P-NMR characteristics of intact rat liver mitochondria, mitoplasts and isolated inner mitochondrial membranes, as well as mitochondrial phosphatidylethanolamine and phosphatidylcholine, have been examined.

2. 2. Rat liver mitochondrial phosphatidylethanolamine hydrated in excess aqueous buffer undergoes a bilayer-to-hexagonal (H_II) polymorphic phase transition as the temperature is increased through 10°C, and thus prefers the H_II) arrangement at 37°C. Rat liver mitochondrial phosphatidylcholine, on the other hand, adopts the bilayer phase at 37°C.

3. 3. Total inner mitochondrial membrane lipids, dispersed in an excess of aqueous buffer, exhibit ³¹P-NMR spectra consistent with a bilayer arrangement for the majority of the endogeneous phospholipids; the remainder exhibit spectra consistent with structure allowing isotropic motional averaging. Addition of Ca²⁺ results in hexagonal (H_II) phase formation for a portion of the phospholipids, as well as formation of ‘lipidic particles’ as detected by freeze-fracture techniques.

4. 4. Preparations of inner mitochondrial membrane at 4 and 37°C exhibit ³¹P-NMR spectra consistent with a bilayer arrangement of the large majority of the endogenous phospholipids which are detected. Approx. 10% of the signal intensity has characteristics indicating isotropic motional averaging processes. Addition of Ca²⁺ results in an increase in the size of this component, which can become the dominant spectral feature.

5. 5. Intact mitochondria, at 4°C, exhibit ³¹P-NMR spectra arising from both phospholipid and small water-soluble molecules (ADP, P_i, etc.). The phospholipid spectrum is characteristic of a bilayer arrangement. At 37°C the phospholipids again give spectra consistent with a bilayer; however, the labile nature of these systems is reflected by increased isotropic motion at longer (at least 30 min) incubation times.

6. 6. It is suggested that the uncoupling action of high Ca²⁺ concentrations on intact mitochondria may be related to a Ca²⁺-induced disruption of the integrity of the inner mitochondrial phospholipid bilayer. Further, the possibility that non-bilayer lipid structures such as inverted micelles occur in the inner mitochondrial membrane cannot be excluded.

Primary structure of mitochondrial glutamic oxaloacetic transaminase from rat liver: Comparison with that of the pig heart isozyme

The complete amino acid sequence of the mitochondrial glutamic oxaloacetic transaminase isozyme from rat liver is presented. The sequence contained 401 amino acid residues, 10 of which are methionine. Cyanogen bromide cleavage of mitochondrial glutamic oxaloacetic transaminase produced 12 peptides, one of which contained an internal homoserine residue resulting from incomplete cleavage by cyanogen bromide. The calculated molecular weight was 44,358. The sequence showed 94% homology with that of the corresponding isozyme from pig heart. These findings support the conclusion that the rate of evolution of the mitochondrial isozymes is lower than that of their cytosolic isozymes.     

An improved method for estimating sequence divergence between related DNAs from changes in restriction endonuclease cleavage sites

Summary We have developed a theory to estimate the degree of sequence divergence between related DNAs from the comparison of restriction endonuclease recognition sites. Two major improvements have been made upon a similar method reported by Upholt (1977). First, the most probable value is calculated by the collective use of all available data. This reduces intrinsic statistical error and extends the analyzable range of sequence divergence. Second, all variables are redefined so that they have strict mathematical implications. This corrects a serious error arising from the misinterpretation of the meaning of the fraction of conserved cleavage sites. With this refined method, sequence divergence between rat and mouse mitochondrial DNAs (mtDNAs) was calculated to be about 25% substitutions/nucleotide, which is in good agreement with the DNA-DNA hybridization data obtained by Jakovcic et al. (1975). It was also estimated that the three types of rat mtDNAs differ from one another by 0.3 ~1% of total base pairs. These values are 2 ~5 times smaller than those obtained with the conventional method.     

Mitochondrial mutagenesis in Saccharomyces cerevisiae. III. Nitrous acid.

Nitrous acid (NA) induced mutations efficiently in mitDNA, conferring resistance to erythromycin and weakly induces mit- mutations. In some strains of yeast it also enhanced rho- mutations. The frequencies of nuclear and mitochondrial mutations induced with NA are compared.     

Precise localization of the origin of replication in a physical map of HeLa cell mitochondrial DNA and isolation of a small fragment that contains it

The precise positions of the origin of replication³ and of the D-loop within the HpaII restriction map of HeLa cell mitochondrial DNA have been investigated. For this purpose, 7 S DNA, which is the heavy-chain initiation sequence, was used as a template for fragment-primed DNA synthesis by Escherichia coli DNA polymerase I. The results indicate clearly that the origin of replication lies in HpaII fragment 8 at about 80 base-pairs from the border with fragment 17, and that the D-loop region extends from this site, through fragment 17, to a position in fragment 10 which is about 365 base-pairs from the border with fragment 17. Sequential digestion of fragment 8 with HaeIII enzyme has allowed the isolation of a subfragment, about 200 base-pairs long, that contains the origin of replication.