DNA polymorphism in the Mongolian population. Restriction fragment length polymorphism analysis of mitochondrial DNA]

Restriction enzyme fragment patterns in the D loop and deletion-insertion polymorphism in the V noncoding region of human mitochondrial DNA (mt DNA) were analysed in Mongolian population using the polymerase chain reaction. Polymorphisms were detected and mt DNAs classified into 40 types using seven enzymes--AvaII, BamHI, CfrI131, KpnI, EcoRV, HaeIII RsaI and Asian specific deletion and insertion. The allele frequencies of the polymorphisms and gene diversity were determined. The data obtained for Mongolian population and the literature data were comparatively studied.     

Different base per unit length ratios exist in single-stranded RNA and single-stranded DNA.

A significant difference was found to exist in the number of bases per unit length of single-stranded RNA as compared to single-stranded DNA when single-stranded RNA or DNA molecules of known nucleotide sequence were measured by electron microscopy using a cytochrome spreading technique. Using this technique, single-stranded RNA was found to have 17.5% more bases per unit of length than single-stranded DNA. These ratios were verified using four different denaturing conditions for the RNA: 80% formamide, 80% formamide plus glyoxal, 80% formamide/4M urea and 80% formamide/4M urea plus glyoxal. Molecules ranging in size from 1541 to 5386 nucleotides were examined and the number of bases per unit length was found to vary inversely with micrometer was consistent when RNA and DNA molecules of the same length were compared.     

DNA fingerprinting of Leonurus cardiaca L. germplasm in Iran using amplified fragment length polymorphism and inter-retrotransposon amplified polymorphism

In the present study, the extent of inter and intra-population genetic variation was evaluated in Leonurus cardiaca accessions naturally growing in Iran by AFLP and IRAP markers. The fingerprints corresponding to AFLP and IRAP markers revealed high levels of heterozygosity, indicating that L. cardiaca is predominantly an out-crossing species. The average percentage polymorphism was detected as 58% and 90.8% on utilizing AFLP and IRAP data, respectively. Gene diversity values within populations varied 0.14 to 0.20 for AFLP and 0.12 to 0.21 for IRAP. The overall levels of genetic variation present in the L. cardiaca germplasm in Iran were finally determined by combining the AFLP and IRAP datasets to ensure wide genome coverage. The phenogram depicted that the accessions of Dargaz population were genetically distinct from other populations. Based on AFLP and IRAP analysis, it is concluded that L. cardiaca maintains high levels of genetic variation at inter and intra-population level.     

Inheritance and restriction fragment length polymorphism of chloroplast DNA in the genus Coffea L.

CpDNA variation among 52 tree samples belonging to 25 different taxa of Coffea and two species of Psilanthus was assessed by RFLP analysis on both the total chloroplast genome and the atpB-rbcL intergenic region. Twelve variable characters were distinguished allowing the identification of 12 different plastomes. The low sequence divergence observed might suggest that Coffea is a young genus. The results were in contradiction with the present classification into two genera. Additionally, cpDNA inheritance was studied in interspecific hybrids between C. arabica and C. canephora, and in an intraspecific progeny of C. canephora, using PCR-based markers. Both studies showed exclusively maternal inheritance of cpDNA.     

DNA polymorphism in the Mongolian population. Restriction fragment length polymorphism analysis of DNA at seven loci of the nuclear genome]

Seven DNA markers from five genes and one chromosomal region were analysed in Mongolian population using the polymerase chain reaction. The frequencies of alleles of the polymorphisms detected with HindIII in the HBG-2, AvaII in the HBB, MspI and XbaI in the Apo-B, PstI in the D7S8, HincII in the LDLR and allele frequency of the minisatellite fragment in the AT-3 have been determined. The results of the RELP for Apo-B(MspI), LDLR, D7S8 and AT-3 are obtained for the first time among Mongoloids. DNA markers studied demonstrated high level of polymorphisms in the population of Mongolia, except for XbaI and MspI restriction sites at the Apo-B locus. The data obtained for Mongolian population and the literature data were compared.     

Rapid DNA purification for restriction fragment length polymorphism analysis

This paper describes a method for isolation of DNA from blood samples involving a rapid chemical disintegration of proteins with 8 M urea and with a minimum of exposure to phenol. The DNA is further desalted and purified on Sephadex G-25 prepacked disposable columns. DNA isolated in this way was pure enough to be immediately cleaved by restriction enzymes.     

Restriction fragment length polymorphism in the mitochondrial DNA of cloned cattle

The clonal origin of 4 Holstein bulls was determined by hybridization experiments with 2 different minisatellite probes, and all 4 animals showed identical genomic DNA fingerprints, hence confirming monozygosity. Extra-chromosomal differences were observed among these 4 Holstein bulls. Mitochondrial DNA restriction fragment length polymorphisms with restriction endonucleases Avall and Hhal sites were found, and these polymorphisms can be explained by the loss of a single site for each of these 2 enzymes. Since mitochondrial DNA are maternally transmitted, all 4 bulls would produce genetically equivalent spermatozoa and offspring. The combination of embryo cloning and specific cytoplasmic markers would provide an ideal system for the study of maternal cytoplasmic effects on quantitative traits.     

Restriction fragment length polymorphism detected by cDNA and genomic DNA clones in Stylosanthes

A DNA isolation method suitable for genomic library construction and RFLP analyses of the forage legume Stylosanthes was developed. Probes isolated using this method were used to investigate the feasibility of constructing RFLP-based genetic maps in this genus. Two hundred and seventy-one PstI genomic DNA and 134 cDNA clones were analysed against four Stylosanthes accessions, including two tetraploids and two diploids, with the use of two restriction enzymes, DraI and HindIII. The proportion of clones which detected single-copy sequences from the PstI genomic library was higher than that from the cDNA library, but the percentage of clones which detected low-copy sequences was doubled in the latter. There was no significant difference in the level of RFLPs detected by gDNA and cDNA probes, although the level of polymorphism was lower in the diploids. A large proportion of RFLPs seemed to have resulted from mutation/base substitution events, and this was especially the case in diploids.     

DNA length polymorphism located 5'' to the human myelin basic protein gene.

A region of DNA 5' to the human myelin basic protein (MBP) gene, located on the long arm of chromosome 18, is a site of restriction-fragment-length polymorphism (RFLP) showing 37% heterozygosity in 40 subjects studied. Southern transfer analysis using a 0.9-kb genomic fragment encompassing the first exon of the human MBP gene reveals this polymorphism with at least nine restriction enzymes, indicating that insertion, deletion, or both is the basis for the DNA length variation. Double restriction-enzyme digest analysis suggests that this polymorphism is within the region 0.5-2.0 kb upstream of the coding region of the first exon of the human MBP gene. Eleven different allelic RFLPs were identified, differing in size by as many as 450 bp. The distribution of insertion/deletion-size variants from this region is bimodal, with most restriction fragments varying in size over a 0.1-kb range. Pedigree analysis of polymorphism at this site in one three-generation family shows Mendelian assortment of parental haplotypes. The form and frequency of polymorphism generated by this site is similar to that reported for human DNA regions comprised of homologous short tandem repeats.     

Potassium transport in two tomato species : lycopersicon esculentum and lycopersicon cheesmanii

The commercial tomato, lycopersicon esculentum Mill. cv Walter, and its wild relative, Lycopersicon cheesmanii ssp. minor (Hook.) C.H. Mull., were grown for 30 days under controlled conditions and in solution culture varying in its content of Na⁺ and K⁺. Subsequently, ⁸⁶Rb-labeled K⁺ uptake and distribution were studied. From all solutions, `Walter' consistently absorbed <sup>86</sup>Rb-labeled K<sup>+</sup> at a higher rate in micromoles per gram fresh weight per 30 minutes than L. cheesmanii. L. cheesmanii distributed a greater proportion of the absorbed K<sup>+</sup> from its root to its shoot. When 0.6 millimolar NaNO<sub>3</sub> replaced 0.6 millimolar KNO<sub>3</sub> in the pretreatment solution, intact plants of both genotypes followed a similar pattern as when they were pretreated with K<sup>+</sup> only, but a greater percentage of the absorbed K<sup>+</sup> remained in the roots. Leaf slices of L. cheesmanii plants deprived of K<sup>+</sup> for 6 days showed a greater rate of K<sup>+</sup> uptake than did slices from `Walter' plants pretreated the same way. Stem slices of L. cheesmanii, however, had a lower uptake rate than did those of `Walter'. Both leaf and stem slices of `Walter' plants, pretreated 6 days with 0.6 millimolar NaNO₃ substituting for 0.6 millimolar KNO₃ in their growth medium, had greater rates of ⁸⁶Rb-labeled K⁺ uptake from 0.5 and 20 millimolar KCl solutions than did slices of L. cheesmanii. These marked differences in patterns of ion uptake and translocation indicate that these genotypes of tomato have evolved different mechanisms to deal with K⁺ and Na⁺ in their environments.     

Amplified fragment length polymorphism markers for DNA fingerprinting in the genus Salvia

Abstract

The interspecific relationships among 51 worldwide collected accessions of Salvia have been investigated using the amplified fragment length polymorphism (AFLP) technique. The assessed genetic similarities allowed us to group the genotypes into two main clusters according to their geographical origin. Our results are encouraging for further characterization of the genus with the aim to clarify Salvia taxonomy.     

DNA variation in the wild plant Arabidopsis thaliana revealed by amplified fragment length polymorphism analysis.

To investigate the level and pattern of DNA variation of Arabidopsis thaliana at the entire genome level, AFLP analysis was conducted for 38 ecotypes distributed throughout the world. Ten pairs of selective primers were used to detect a total of 472 bands, of which 374 (79. 2%) were polymorphic. The frequency distribution of polymorphic bands was skewed toward an excess of singleton variation. On the basis of AFLP variation, nucleotide diversity for the entire genome was estimated to be 0.0106, which was within the range reported previously for specific nuclear genes. The frequency distribution of pairwise distance was bimodal because of an ecotype (Fl-3) with a large number of unique bands. Linkage disequilibrium between polymorphic AFLPs was tested. The proportion of significant linkage disequilibria was close to random expectation after neglecting the ecotype Fl-3. This result indicates that the effect of recombination could not be ignored in this selfing species. A neighbor-joining tree was constructed on the basis of the AFLP variation. This tree has a star-like topology and shows no clear association between ecotype and geographic origin, suggesting a recent spread of this plant species and limited migration between its habitats.     

A DNA polymorphism in close physical linkage with the proopiomelanocortin gene

Cellular DNAs from a panel of 20 unrelated individuals were screened for restriction fragment length polymorphisms (RFLP) with a DNA probe containing the first exon of the proopiomelanocortin gene (POMC), which has been assigned to chromosome 2p23-25. Digestion with the restriction endonuclease Sst 1 revealed a high frequency RFLP. The two alleles that were found are fragments of 10- and 15-kilobase (kb) length and are in Hardy-Weinberg equilibrium with frequencies of 72.6% and 27.4%, respectively. Informative families were tested for linkage between POMC/Sst 1 RFLP and other polymorphic markers of chromosome 2. Linkage was excluded to AcP-1 (2p23-25) at 15% recombination, which is still consistent with the chromosomal assignments for these genes. The close physical linkage (10 kb) of the polymorphic locus to the POMC gene makes this RFLP a suitable marker for future linkage studies involving the POMC gene.     

DNA polymorphism in the Russian population. Analysis of dna restriction fragment length polymorphism in seven loci of the nuclear genome]

Using the method for polymerase chain reaction the polymorphism of eight markers of the nuclear DNA was studied. In a sample of Russians taken at random (N = 118) from predominantly southern and central regions of Russia, allele frequencies were determined for restriction sites HindIII at HBG-2 and PAH loci, AvaII at the HBB locus, MspI at the ApoB locus, PstI at D7S8, HincII at LDLR, TaqI and BamHI at the DSX164. Comparative data for different world regions are presented.     

DNA polymorphism in cytokine genes based on length variation in simple-sequence tandem repeats

The possibility of the involvement of cytokines in the genetic predisposition to various diseases has been suggested by a large variety of studies. However, the study of potential disease linkage of cytokine genes has been hampered by a lack of sufficiently polymorphic markers at the restriction fragment length polymorphism (RFLP) level. We have investigated the distribution, the length polymorphism, the informativeness, and the efficiency of analysis, of simple-sequence tandem repeats in the mouse cytokine genes. Highly polymorphic sequences have been identified in the IL-1, IL-1ra, IL-2, IL-4, IL-6, IL-7, and IFN- genes. The utility and the value of these sequences as gene markers is exemplified by mapping the IL-7 gene to mouse chromosome 3 close to pgk-1ps3 and Car-2 loci and the IFN- gene to chrrmosome 10 near the pg locus. Advantages of short tandemly repeated sequences as genetic markers are discussed in comparison with RFLPs.     

Small angle x-ray scattering of chromatin. Radius and mass per unit length depend on linker length.

Analyses of low angle x-ray scattering from chromatin, isolated by identical procedures but from different species, indicate that fiber diameter and number of nucleosomes per unit length increase with the amount of nucleosome linker DNA. Experiments were conducted at physiological ionic strength to obtain parameters reflecting the structure most likely present in living cells. Guinier analyses were performed on scattering from solutions of soluble chromatin from Necturus maculosus erythrocytes (linker length 48 bp), chicken erythrocytes (linker length 64 bp), and Thyone briareus sperm (linker length 87 bp). The results were extrapolated to infinite dilution to eliminate interparticle contributions to the scattering. Cross-sectional radii of gyration were found to be 10.9 +/- 0.5, 12.1 +/- 0.4, and 15.9 +/- 0.5 nm for Necturus, chicken, and Thyone chromatin, respectively, which are consistent with fiber diameters of 30.8, 34.2, and 45.0 nm. Mass per unit lengths were found to be 6.9 +/- 0.5, 8.3 +/- 0.6, and 11.8 +/- 1.4 nucleosomes per 10 nm for Necturus, chicken, and Thyone chromatin, respectively. The geometrical consequences of the experimental mass per unit lengths and radii of gyration are consistent with a conserved interaction among nucleosomes. Cross-linking agents were found to have little effect on fiber external geometry, but significant effect on internal structure. The absolute values of fiber diameter and mass per unit length, and their dependencies upon linker length agree with the predictions of the double-helical crossed-linker model. A compilation of all published x-ray scattering data from the last decade indicates that the relationship between chromatin structure and linker length is consistent with data obtained by other investigators.     

TheGunnera symbiosis: DNA restriction fragment length polymorphism and protein comparisons ofNostoc symbionts

Cyanobacteria separated from symbiosis with several species of the angiospermGunnera were comparatively characterized and correlated with the locales and taxonomy of their host plants. All were identified as strains ofNostoc. Protein profiles and DNA restriction fragment length polymorphisms (from hybridizations with heterologousnifH andglnA probes) determined that three of the four cyanobacteria fromGunnera grown at one site in Sweden, each from a different host species, were very similar or identical. Plants of one species,G. manicata, grown in a second location at the site were infected with a different cyanobiont. Among five isolates from two species ofGunnera, collected in the same locale in New Zealand, three subgroups were documented. Isolates from three differentGunnera species grown in separate locations in the United States were each uniquely different. None of the cyanobacteria differed in the molecular weights of their glutamine synthetase and Fe-nitrogenase proteins. The diversity and accessibility of compatibleNostoc populations present in the soil micro-environment, not a critical selective factor required byGunnera, were concluded to be a major determinant in symbiont selection.     

Potassium and sodium absorption kinetics in roots of two tomato species : lycopersicon esculentum and lycopersicon cheesmanii

Excised roots of the tomato species, Lycopersicon esculentum Mill. cv Walter (the commercial species) and of Lycopersicon cheesmanii ssp. minor (Hook.) C.H. Mull. (a wild species from the Galapagos Islands), were used in comparative studies of their absorption of K⁺ and Na⁺. Uptake of ⁸⁶Rb-labeled K⁺ and ²²Na-labeled Na⁺ by excised roots of `Walter' and L. cheesmanii varied as a function of genotype and tissue pretreatment with or without K<sup>+</sup>. Excised roots of `Walter' consistently absorbed more ⁸⁶Rb-labeled K⁺ than those of L. cheesmanii. Absorption of K⁺ from solutions ranging from 0.01 to 0.2 millimolar KCl showed saturation kinetics in both K⁺-pretreated and K⁺-depleted roots of `Walter,' and for K<sup>+</sup>-depleted roots of L. cheesmanii. K<sup>+</sup>-pretreated roots of L. cheesmanii had exceedingly low rates of K<sup>+</sup> uptake with strikingly different, linear kinetics. Pretreatment with K<sup>+</sup> caused a decrease in rates of K<sup>+</sup> uptake in both genotypes. Potassium depleted roots of L. cheesmanii absorbed Na<sup>+</sup> at a greater rate than those of `Walter,' whereas K⁺-pretreated roots of `Walter' absorbed Na⁺ at a greater rate than those of L. cheesmanii. The results confirm and extend previous conclusions to the effect that closely related genotypes may exhibit widely different responses to the two alkali cations, K⁺ and Na⁺.