Increase In Amount Of DNA Research Articles

Evolution of Genome Size within the Genus Haliotis (Vetigastropoda: Haliotidae)

Genome size (C-value) and the percentage of adenine and thymine nucleotides in the genome (AT content) are fundamental characteristics of every species, and very important parameters in molecular cytogenetic and phylogenic studies, and for the progress of whole-genome sequencing. In this study, the C-value and AT content of nine abalone species inhabiting the Northwest Pacific (around Japan), Southeast Asia, South Africa, and Oceania were determined using flow cytometry. C-value ranged from 1.32 pg for Haliotis varia (Southeast Asia) to 2.01 pg for Haliotis laevigata (Oceania), and AT content ranged from 58.0% for Haliotis planata (Southeast Asia) to 66.3% for Haliotis diversicolor aquatilis (Southeast Asia). Reported chromosome numbers ranged from 2n = 32 to 36, and were higher in Oceanian, South African, and Northwest Pacific species (2n = 36) compared with the Southeast Asian group (2n = 32). This increase of chromosome number in the Oceanian species seems to be related to an increase in the DNA amount, unlike that of Haliotis midae (South African species). An increase of the C-value associated with an increase in DNA amount was also shown in the North Pacific group. These results suggest that increases in the DNA amount occurred independently in the ancestor of each of the different lineages.

Purmorphamine increased adhesion, proliferation and expression of osteoblast phenotype markers of human dental pulp stem cells cultured on beta-tricalcium phosphate.

Growth factors play a significant role in cell proliferation and differentiation during different stages of the bone repair. However, several limitations have been brought researchers attention to an osteoinductive small molecule including Purmorphamine. In this study, we aimed to evaluate the effect of Purmorphamine on adhesion, proliferation and differentiation of human dental pulp stem cells (hDPSCs) seaded on beta-tricalcium phosphate (β-TCP) granules. hDPSCs were established from extracted wisdom teeth of healthy volenteers. Cells at passage 3 were seeded on β-TCP in the presence or absence of Purmorphamine. Cell adhesion and proliferation were assessed using scanning electeron microscopy (SEM) and DNA counting assay, respectively, after 1, 3 and 5days. Then, hDPSCs seeded on β-TCP were subjected to osteogenic medium with or without Purmorphamine. After 7 and 14days osteogenic diffrentiation capability of hDPSCs were determined using real-time RT-PCR and alkaline phosphatase (ALP) activity assay. The significant increase in amount of DNA was observed at day 3 and 5 in the presence of Purmorphamine. SEM imaging also was confirmed the DNA counting assay; in all given time points, hDPSC attachment and growth was significantly higher in the presence of Purmorphamine. ALP activity was increased by Purmorphamine at both 7 and 14days of induction. Purmorphamine showed to effect on osteopontin expression at earlier stage of osteogenic differentiation, whereas for osteocalcin expression, this effect was more evident at later stage of differentiation. Purmorphamine had a promotive effect on adhesion, proliferation and osteogenic differentiation of hDPSCs cultured on β-TCP. The outcome of the current study would help in development of in vitro culture conditions for better osteogenic differentiation of hDPSCs prior to transplantation.

Specific activity of thymidine nucleotide biosynthetic enzymes in hairy roots extracts of Sesamum indicum L. transformed by two strains of Agrobacteruim rhizogenes

The study concluded induction of transformed hairy roots from leaves and decapitated seedlings of Sesamum indicum L. using two strains of Agrobacterium rhizogenes considered as natural vector of transformation. The strain R1601 stimulated roots on leaves and seedlings during 20 days of inoculation placed on solidified Arnon and Hoagland medium. Whereas they involved 12 days when inoculated with the strain R15834. Generally strain R15834 was efficient in inducing these roots and their numbers than strain R1601 which approached 54.4% and 41.6% respectively. The results indicated an increase in the specific activity of enzymes Thymidlate synthase (TS), Dihydrofolate reductase (DHFR), Serine hydroxy methyl transfrase (SHMT) in extract of transformed hairy roots producing agropine by stain R15834 and approach 4.610, 1.057, 0.480 µmolmin mg of protein respectively compared with the activity of 1.256, 0.097, 0.125µ molmin mg in the control samples. This was coupled with an increase in amount of DNA and RNA that approached 105, 1020 µg per gram respectively compared to 44, 462 µg per gram in control samples. The transformation of these hairy roots was pointed out through the separation of agropine spots from their extracts when electrophoreted in the presence of standared agropine.

Biochemical investigation of yttrium(III) complex containing 1,10-phenanthroline: DNA binding and antibacterial activity

Characterization of the interaction between yttrium(III) complex containing 1,10-phenanthroline as ligand, [Y(phen)2Cl(OH2)3]Cl2⋅H2O, and DNA has been carried out by UV absorption, fluorescence spectra and viscosity measurements in order to investigate binding mode. The experimental results indicate that the yttrium(III) complex binds to DNA and absorption is decreasing in charge transfer band with the increase in amount of DNA. The binding constant (Kb) at different temperatures as well as thermodynamic parameters, enthalpy change (ΔH°) and entropy change (ΔS°), were calculated according to relevant fluorescent data and Vant’ Hoff equation. The results of interaction mechanism studies, suggested that groove binding plays a major role in the binding of the complex and DNA. The activity of yttrium(III) complex against some bacteria was tested and antimicrobial screening tests shown growth inhibitory activity in the presence of yttrium(III) complex.

Gene delivery to human bone marrow mesenchymal stem cells by microporation

Electroporation has been considered one of the most efficient non-viral based methods to deliver genes regardless of frequently observed high cell mortality. In this study we used a microporation technique to optimise the delivery of plasmid DNA encoding green fluorescence protein (GFP) to human bone marrow mesenchymal stem cells (BM-MSC). Using resuspension buffer (RB) and as low as 1.5 × 10 5 cells and 1 μg of DNA, we achieved 40% of cells expressing the transgene, with cell recovery and cell viabilities of 85% and 90%, respectively. An increase in DNA amount did not significantly increase the number of transfected cells but clearly reduced cell recovery. A face-centered composite design was used to unveil the conditions giving rise to optimal plasmid delivery efficiencies when using a sucrose based microporation buffer (SBB). The BM-MSC proliferation kinetics were mainly affected by the presence of plasmid and not due to the microporation process itself although no effect was observed on their immunophenotypic characteristics and differentiative potential. Based on the data shown herein microporation demonstrated to be a reliable and efficient method to genetically modify hard-to-transfect cells giving rise to the highest levels of cell survival reported so far along with superior gene delivery efficiencies.

IGF-I- and EGF-dependent DNA synthesis of porcine myoblasts is influenced by the dietary isoflavones genistein and daidzein

Soy-derived isoflavones have been reported to be specific inhibitors of protein tyrosine kinases like the type 1 insulin-like growth factor receptor (IGF-1R) and the epidermal growth factor receptor (EGFR). This study was conducted to investigate, whether IGF-I and EGF stimulate porcine myoblast growth and whether the responses are influenced by isoflavones. Satellite cell-born myoblasts derived from the semimembranosus muscle of newborn piglets were treated for 26 h with IGF-I or EGF alone and in combination with genistein or daidzein. The DNA amount was measured and DNA synthesis was recorded as 6 h-[ 3H]thymidine incorporation during exponential growth in serum-free basal medium. IGF-I and EGF synergistically stimulated DNA synthesis of porcine myoblast with EGF causing a greater response. Genistein (100 μmol/l) effectively reduced the growth factor-mediated DNA synthesis, which was associated with an inhibition of growth factor receptor protein expression. In response to daidzein no reduction in growth factor-mediated DNA synthesis was found. Daidzein (1; 10 μmol/l) combined with IGF-I caused even a slight increase in DNA amount compared with the untreated control. The expression of the IGF-1R precursor protein was reduced with 10 and 100 μmol/l daidzein, whereas the EGFR expression remained unchanged with daidzein. The results suggest that dietary isoflavones may interact with growth factor-induced stimulation of pig skeletal muscle growth.

DNA-binding proteins of mammalian mitochondria

Acid-soluble proteins were isolated from liver and spleen mitochondria and their ability to form complexes with DNA was investigated. According to electrophoresis data, acid-soluble proteins include about 20 polypeptides ranging in the molecular mass from 10 to 120 kDa. It was found that acid-soluble proteins form stable DNA–protein complexes at a physiological NaCl concentration. Different polypeptides possess different degrees of DNA affinity. There is no significant difference between DNA-binding proteins of mitochondria from liver and those from spleen as to their ability to form complexes with mtDNA and nDNA. In the presence of 5 μg of DNA most polypeptides were bound to DNA, and further increase in DNA amount affected little the binding of proteins to DNA. There was no distinct difference in DNA–protein complex formation of liver mitochondrial acid-soluble proteins with nDNA or mtDNA. Also, it was detected that with these mitochondrial acid-soluble proteins, proteases that specifically cleave these proteins are associated. It was shown for the first time that these proteases are activated by DNA. DNA-binding proteins including DNA-activated mitochondrial proteases are likely to participate in the regulation of the structural organization and functional activity of mitochondrial DNA.

Microvascular development and growth of uterine tissue during the estrous cycle in mares.

To document uterine growth and microvascular development in the endometrium of uteri with differing degrees of fibrosis as well as uterine growth throughout the estrous cycle of mares. 30 mares. Uterine tissue was obtained during the breeding season from a slaughter facility. Stage of estrous cycle of the mares was assessed on the basis of ovarian structures and plasma progesterone concentrations. Endometrium was characterized by use of light microscopy, and blood vessel walls were marked by histochemical techniques. Microvascular development was evaluated by a computerized image analysis system. Growth of uterine tissue was based on cellular content of DNA and RNA, RNA:DNA, and protein:DNA. Significant differences in vascular density were not observed in the endometrium of uteri obtained from mares euthanatized during the follicular or luteal phase of the estrous cycle, regardless of whether endometrial classification of degree of fibrosis was considered. There was a 3-fold increase in amount of DNA and RNA of endometrial cells in the follicular phase when compared to myometrium. Hypertrophy of endometrial tissue during the luteal phase was reflected by a significant increase in cell protein content and protein:DNA. Endometrial growth of vascular tissues during the estrous cycle may be coordinated with development of nonvascular tissue. Estrogen and progesterone may play a role in regulation of uterine growth and angiogenesis.

Genome Size Variation in the Aloaceae, an Angiosperm Family Displaying Karyotypic Orthoselection

Chromosomal variation is widespread in angiosperms, with numbers ranging from 2 n=4 in some species to over 600 in others, and with genome sizes from the minute in Arabidopsis thalianato the very large in Fritillariaspp. Many families show extensive karyotype variation, sometimes within a species, often with a genus. Others display karyotypic orthoselection, with identical karyotypes throughout a genus, or in groups of genera, e.g. Agaveand its relatives. The highest orthoselection level occurs in the Aloaceae, which has the same bimodal karyotype in all species ( x=7; four long and three short acrocentrics). Major structural changes to Aloaceae chromosomes are common in the wild, but are not known as homozygotes and take no part in speciation. The ratio of combined volumes of the long and short chromosomes is constant in Aloespecies. Despite the karyotypic uniformity, 4C nuclear DNA amounts vary two-fold in diploid species, with lowest values in primitive species. This range is demonstrable in the extreme heteromorphy of homeologues that is visible in hybrids. Supported by the observation that chiasma position in an intergeneric hybrid is random, even where homeologue length differs greatly, these findings suggest that evolutionary increase in DNA amount involves the amplification of many small DNA segments at sites distributed evenly throughout the chromosome complement in numbers proportional to chromosome length, thus maintaining karyotypic uniformity in the genus.

Heterochromatin and repetitive DNA frequency variation in regenerated plants of Helianthus annuus L.

Plant regeneration from cotyledons of seeds of a single progeny of a pure line of Helianthus annuus was studied in respect of the nuclear DNA contents of control and regenerated plants. Control plants were divided into two groups: those developed from seeds at the periphery of the inflorescence (showing a high basic 4C DNA content) and those from seeds developed in the middle of the inflorescence (showing a low basic 4C DNA content). It was observed that plants from peripheral seeds have a higher morphogenetic potential than those from central seeds. Cytophotometric analyses indicated that plants regenerated from cotyledons of both peripheral and central seeds show the same basic 4C DNA amount, which is higher that that observed in vivo in peripheral seeds. Molecular analysis by slot blotting and hybridization with different DNA families showed that the difference in nuclear DNA content between plants from peripheral and central seeds in vivo are mainly related to differences in the frequency of highly repeated, "slow" medium repeated (MR2), and ribosomal DNA families; by contrast, the increase in DNA amount in regenerated plants is mainly due to "fast" medium repeated sequences (MR1). Moreover, the frequency of kinetically isolated "unique" sequences was higher in peripheral seeds than in central ones and still higher in regenerated plants. Optical-density measurements of interphase nuclei showed an increase of heterochromatin in regenerated plants, suggesting that, whatever DNA is amplified in these plants, it remains condensed and probably inactive.

Quantitative nuclear DNA changes inEleusine (Gramineae)

2C nuclear DNA amounts were determined in 30 collections belonging to 10 species ofEleusine. About a 2.5-fold variation in genome size is evident in the genus. The 2C DNA amount in the diploid species ranged from 2.50 pg inE. verticillata to 3.35 pg inE. intermedia. In contrast, the tetraploid species showed a range from 4.95 pg inE. africana to 6.13 pg inE. floccifolia. At intraspecific level 10 collections ofE. coracana, 6 ofE. indica, 4 ofE. africana, 2 ofE. tristachya, and 2 ofE. kigeziensis did not show any significant variation. However, 2 collections ofE. floccifolia, connected with polyploidy, displayed about 90% variation. Polyploid species showed approximately double the genome size of that of their corresponding diploids. An evolutionary increase in DNA amount is evident inE. coracana during the course of its origin and domestication fromE. africana.

Variation in nuclear DNA in the genus Secale

Estimates of the 4C DNA amount per nucleus in 16 taxa of the genus Secale made by Feulgen microdensitometry ranged from 28.85 picograms (pg) in S. silvestre PBI R52 to 34.58 pg in S. vavilovii UM 2D49, compared with 33.14 pg in S. cereale cv. “Petkus Spring” which was used as a standard. Giemsa C-banding patterns showed considerable interspecific and intraspecific variation and several instances of polymorphism for large telomeric C-bands. The proportion of telomeric heterochromatin in the genome ranged from about 6% in S. silvestre and S. africanum to about 12% in cultivated rye. A detailed comparison of nine taxa showed no overall relationship between 4C DNA amount and the proportion of telomeric heterochromatin in the genome. However, evidence is presented which strongly supports the notion that the major evolutionary change in chromosome structure in Secale has involved the addition of heterochromatin at, or close to, the telomeres. It is suggested that saltatory amplification events at telomeres were initially responsible for each large increase in DNA amount. Subsequently unequal crossing over between homologues may have played an important secondary role by extending the range of variation in the amount of heterochromatin at a given telomere, while crossing over between non-homologues may have provided a useful mechanism allowing an increase in the DNA amount at one telomere to be distributed between chromosomes.