836,492 research outputs found
Breathing dynamics in heteropolymer DNA
While the statistical mechanical description of DNA has a long tradition,
renewed interest in DNA melting from a physics perspective is nourished by
measurements of the fluctuation dynamics of local denaturation bubbles by
single molecule spectroscopy. The dynamical opening of DNA bubbles (DNA
breathing) is supposedly crucial for biological functioning during, for
instance, transcription initiation and DNA's interaction with selectively
single-stranded DNA binding proteins. Motivated by this, we consider the bubble
breathing dynamics in a heteropolymer DNA based on a (2+1)-variable master
equation and complementary stochastic Gillespie simulations, providing the
bubble size and the position of the bubble along the sequence as a function of
time. We utilize new experimental data that independently obtain stacking and
hydrogen bonding contributions to DNA stability. We calculate the spectrum of
relaxation times and the experimentally measurable autocorrelation function of
a fluorophore-quencher tagged base-pair, and demonstrate good agreement with
fluorescence correlation experiments. A significant dependence of opening
probability and waiting time between bubble events on the local DNA sequence is
revealed and quantified for a promoter sequence of the T7 phage. The strong
dependence on sequence, temperature and salt concentration for the breathing
dynamics of DNA found here points at a good potential for nanosensing
applications by utilizing short fluorophore-quencher dressed DNA constructs.Comment: 11 pages, 8 figure
Intramolecular integration within Moloney murine leukemia virus DNA
By screening a library of unintegrated, circular Moloney murine leukemia virus (M-MuLV) DNA cloned in lambda phage, we found that approximately 20% of the M-MuLV DNA inserts contained internal sequence deletions or inversions. Restriction enzyme mapping demonstrated tht the deleted segments frequently abutted a long terminal repeat (LTR) sequence, whereas the inverted segments were usually flanked by LTR sequences, suggesting that many of the variants arose as a consequence of M-MuLV DNA molecules integrating within their own DNA. Nucleotide sequencing also suggested that most of the variant inserts were generated by autointegration. One of the recombinant M-MuLV DNA inserts contained a large inverted repeat of a unique M-MuLV sequence abutting an LTR. This molecule was shown by nucleotide sequencing to have arisen by an M-MuLV DNA Molecule integrating within a second M-MuLV DNA molecule before cloning. The autointegrated M-MuLV DNA had generally lost two base pairs from the LTR sequence at each junction with target site DNA, whereas a four-base-pair direct repeat of target site DNA flanked the integrated viral DNA. Nucleotide sequencing of preintegration target site DNA showed that this four-base-pair direct repeat was present only once before integration and was thus reiterated by the integration event. The results obtained from the autointegrated clones were supported by nucleotide sequencing of the host-virus junction of two cloned M-MuLV integrated proviruses obtained from infected rat cells. Detailed analysis of the different unique target site sequences revealed no obvious common features
Position Effect Takes Precedence Over Target Sequence in Determination of Adenine Methylation Patterns in the Nuclear Genome of a Eukaryote, \u3cem\u3eTetrahymena thermophila\u3c/em\u3e
Approximately 0.8% of the adenine residues in the macronuclear DNA of the ciliated protozoan Tetrahymena thermophila are modified to N6-methyladenine. DNA methylation is site specific and the pattern of methylation is constant between clonal cell lines. In vivo, modification of adenine residues appears to occur exclusively in the sequence 5′-NAT-3′, but no consensus sequence for modified sites has been found. In this study, DNA fragments containing a site that is uniformly methylated on the 50 copies of the macronuclear chromosome were cloned into the extrachromosomal rDNA. In the novel location on the rDNA minichromosome, the site was unmethylated. The result was the same whether the sequences were introduced in a methylated or unmethylated state and regardless of the orientation of the sequence with respect to the origin of DNA replication. The data show that sequence is insufficient to account for site-specific methylation in Tetrahymena and argue that other factors determine the pattern of DNA methylation
- …