Role of carcinogen-modified deoxynucleotide precursors in mutagenesis

Agents which damage or modify cellular DNA will generally also modify the nucleotide precursor pools, sometimes preferentially (Topal and Baker, 1982). There are at least two different ways that incorporation of modified (possibly promutagenic) nucleotides could, theoretically, make a significant contribution to the mutations induced by these agents. Modified bases may exhibit ambiguous base pairing and produce mutations during normal replication or they may induce secondary mutations as a result of processing subsequent to incorporation. There are important precedents for such possibilities. Classical studies on mutagenesis with prototype mutagens like 2-aminopurine (2-AP) and 5-bromouracil clearly show that mutations can occur by incorporation of deoxynucleotides of tautomeric or ionized (Sowers et al., 1987) bases into newly synthesized DNA (Ronen, 1979; Lasken and Goodman, 1984, Coulondre and Miller, 1977). 5-Hydroxymethyl-2′-deoxyuridine (HMdU), a product of oxidative DNA damage, can also be (re)incorporated into cellular DNA with both toxic and mutagenic consequences (Kaufman, 1987; Shirname-More et al., 1987). Furthermore, modified nucleotides may alter the pool sizes of the normal nucleotides and indirectly produce toxic and mutagenic effects. However, these effects are generally see at high, nonphysiological, concentrations of the modified precursors and may not be relevant under physiological conditions. The relative importance of modified deoxynucleotide precursors in the production of mutations by alkylating and oxidative DNA-damaging agents is discussed.     

Use of Rolling Circle Amplification to Rapidly Identify Species of Cladophialophora Potentially Causing Human Infection

The genus Cladophialophora comprises etiologic agents of disease in immunocompetent patients, ranging from mild cutaneous colonization to cerebral encephalitis, in addition to saprobic species. Due to the high degree of phenotypic similarity between closely related species of the genus, identification problems are imminent. In the present study, we described rapid and sensitive rolling circle amplification (RCA) method based on species-specific padlock probes targeted for the internal transcribed spacer regions of rDNA. ITS regions of 12 Cladophialophora species were sequenced, and subsequently, 10 specific padlock probes were designed for the detection of single nucleotide polymorphisms. The majority of circularizable padlock probes were designed based on single nucleotide polymorphisms (SNPs), while for C. bantiana, C. immunda and C. devriesii were characterized by two or more nucleotides. Individual species-specific probes correctly identified in all ten Cladophialophora species correctly by visualization on 1.2 % agarose gels used to verify specificity of probe-template binding; no cross-reactivity was observed. Simplicity, sensitivity, robustness and low costs provide RCA a distinct place among isothermal techniques for DNA diagnostics. However, restriction and specificity and sensitivity should be lowered and increased, respectively, to be useful for a wide variety of clinical applications.     

In vitro efficacy of plant essential oils against Phomopsis azadirachtae – the causative agent of die-back disease of neem

Seven essential oils – Rosemary, Wintergreen, Teatree, Patchouli, Palmarosa, Geranium and Eucalyptus were used to determine the in vitro antifungal activity on Phomopsis azadirachtae. This organism is responsible for the destructive die-back disease of neem. The oils mentioned above were screened using the food poisoning technique. All the seven oils showed considerable antifungal activity against P. azadirachtae. After the conduction of the tests using the oils, the results were such that Wintergreen, Geranium and Palmarosa showed complete or 100% inhibition of mycelial growth at a concentration of 2500 ppm. Among the oils tried, Palmarosa proved to be most potent showing complete inhibition of mycelia at 500 ppm. This really suggests that plant sources possess antifungal activity and can be effectively used to treat and manage plant diseases thereby circumventing the fungal spread in our environment.     

Calcium-doped single-wall nanotubes (Ca/SWCNTs) as a superior carrier for atropine drug delivery: a quantum-chemical study in gas and solvent phases

Communicated by Ramaswamy H. Sarma     

Pair-preferences: a Quantitative Measure of Regularities in Protein Sequences

Abstract

We present here the results obtained by applying several different methods to quantitatively measure regularities in protein sequences based on pair-preferences. We have studied the distribution of amino acid residues, singly as well as in pairs in a large data base and have attempted this task. We confirmed the existence of well-defined pair-preferences in proteins which were shown to be remarkably absent in simulated random sequences of similar amino acid distribution. The analysis of the sequences from the SWISS-PROT data base using simple statistical tests, Fourier analysis, fractal analysis and statistical thermodynamical tests were used to derive parameters to define a natural sequence. As a consequence of the existence of pair-preferences, parameters like fractal dimension (D), spectral exponent (β), scaling parameter (H) and entropy (statistical) were found to be characteristic for natural sequences. For a reference state we chose a randomised state devoid of any pair-preference. The pair-preferences qualified well to be used as quantitative measures of regularities in protein sequences.     

Comparison of SSR and SNP Markers in Estimation of Genetic Diversity and Population Structure of Indian Rice Varieties

Simple sequence repeat (SSR) and Single Nucleotide Polymorphic (SNP), the two most robust markers for identifying rice varieties were compared for assessment of genetic diversity and population structure. Total 375 varieties of rice from various regions of India archived at the Indian National GeneBank, NBPGR, New Delhi, were analyzed using thirty six genetic markers, each of hypervariable SSR (HvSSR) and SNP which were distributed across 12 rice chromosomes. A total of 80 alleles were amplified with the SSR markers with an average of 2.22 alleles per locus whereas, 72 alleles were amplified with SNP markers. Polymorphic information content (PIC) values for HvSSR ranged from 0.04 to 0.5 with an average of 0.25. In the case of SNP markers, PIC values ranged from 0.03 to 0.37 with an average of 0.23. Genetic relatedness among the varieties was studied; utilizing an unrooted tree all the genotypes were grouped into three major clusters with both SSR and SNP markers. Analysis of molecular variance (AMOVA) indicated that maximum diversity was partitioned between and within individual level but not between populations. Principal coordinate analysis (PCoA) with SSR markers showed that genotypes were uniformly distributed across the two axes with 13.33% of cumulative variation whereas, in case of SNP markers varieties were grouped into three broad groups across two axes with 45.20% of cumulative variation. Population structure were tested using K values from 1 to 20, but there was no clear population structure, therefore Ln(PD) derived Δk was plotted against the K to determine the number of populations. In case of SSR maximum Δk was at K=5 whereas, in case of SNP maximum Δk was found at K=15, suggesting that resolution of population was higher with SNP markers, but SSR were more efficient for diversity analysis.     

Providing Spark and Stability: The Role of Intermediary Organizations in Establishing School-Based Youth-Adult Partnerships

The term “youth-adult partnership” increasingly is being used to define a relationship in which both youth and adults have the potential to contribute to decision making processes, to learn from one another, and to promote change. Establishing a relationship with an intermediary organization can provide much needed stability and sustainability to youth-adult partnerships–especially in school settings. Rather than creating generalizable findings, this article focuses on hypothesis generation regarding the ways in which an intermediary organization can provide the needed training and support to school-based youth-adult partnerships. Using a combination of case-study and survey data, the study finds that youth-adult partnerships need help with: youth leadership skill development, team building, project development, and targeted training for adults. Technical assistance needs vary depending on whether groups need to create a spark that will build the idea of a youth-adult partnership or to create stability for a youth-adult partnership that has already been developed     

An alternate protocol for establishment of primary caprine fetal myoblast cell culture: an in vitro model for muscle growth study

Cultured myoblasts have been used extensively as an in vitro model in understanding the underlying mechanisms of myogenesis. Various protocols for establishing a pure myoblast culture have been reported which involve the use of special procedures like flow cytometry and density gradient centrifugation. In goat, only a few protocols for establishing a myogenic cell culture are available and these protocols use adult muscle tissues which often does not yield sufficient numbers of precursor cells with adequate proliferative capacity. Considering the disadvantages of adult myoblasts, we are proposing an alternate protocol using caprine fetus which does not require any special procedures. In the present study, more than 90–95% fetal-derived cell populations had the typical spindle to polyhedral shape of myoblast cell and stained positive for desmin, hence confirming their myogenic origin. These cells attained the maximum confluency as early as 3–4 d against 3 wk by adult myoblasts indicating a better growth potential. Further, quantitative real-time PCR analysis revealed a higher expression (p?<?0.01) of myogenic regulatory factors (i.e., myogenic determination factor 1, myogenic factor 5, and myogenin) and myostatin (MSTN) in the fetal as compared to the adult myoblasts. Consequently, higher proliferation and differentiation ability along with higher abundance of myogenic markers and MSTN make the fetal myoblasts a better in vitro model.     

A Genomics Approach to Deciphering Lignin Biosynthesis in Switchgrass

It is necessary to overcome recalcitrance of the biomass to saccharification (sugar release) to make switchgrass (Panicum virgatum) economically viable as a feedstock for liquid biofuels. Lignin content correlates negatively with sugar release efficiency in switchgrass, but selecting the right gene candidates for engineering lignin biosynthesis in this tetraploid outcrossing species is not straightforward. To assist this endeavor, we have used an inducible switchgrass cell suspension system for studying lignin biosynthesis in response to exogenous brassinolide. By applying a combination of protein sequence phylogeny with whole-genome microarray analyses of induced cell cultures and developing stem internode sections, we have generated a list of candidate monolignol biosynthetic genes for switchgrass. Several genes that were strongly supported through our bioinformatics analysis as involved in lignin biosynthesis were confirmed by gene silencing studies, in which lignin levels were reduced as a result of targeting a single gene. However, candidate genes encoding enzymes involved in the early steps of the currently accepted monolignol biosynthesis pathway in dicots may have functionally redundant paralogues in switchgrass and therefore require further evaluation. This work provides a blueprint and resources for the systematic genome-wide study of the monolignol pathway in switchgrass, as well as other C4 monocot species.