154,890 research outputs found
Human-chimpanzee alignment: Ortholog Exponentials and Paralog Power Laws
Genomic subsequences conserved between closely related species such as human
and chimpanzee exhibit an exponential length distribution, in contrast to the
algebraic length distribution observed for sequences shared between distantly
related genomes. We find that the former exponential can be further decomposed
into an exponential component primarily composed of orthologous sequences, and
a truncated algebraic component primarily composed of paralogous sequences.Comment: Main text: 31 pages, 13 figures, 1 table; Supplementary materials: 9
pages, 9 figures, 1 tabl
An efficient one-step site-directed deletion, insertion, single and multiple-site plasmid mutagenesis protocol
Background: Mutagenesis plays an essential role in molecular biology and biochemistry. It has also been used in enzymology and protein science to generate proteins which are more tractable for biophysical techniques. The ability to quickly and specifically mutate a residue(s) in protein is important for mechanistic and functional studies. Although many site-directed mutagenesis methods have been developed, a simple, quick and multi-applicable method is still desirable. Results: We have developed a site-directed plasmid mutagenesis protocol that preserved the simple one step procedure of the QuikChange (TM) site-directed mutagenesis but enhanced its efficiency and extended its capability for multi-site mutagenesis. This modified protocol used a new primer design that promoted primer-template annealing by eliminating primer dimerization and also permitted the newly synthesized DNA to be used as the template in subsequent amplification cycles. These two factors we believe are the main reasons for the enhanced amplification efficiency and for its applications in multi-site mutagenesis. Conclusion: Our modified protocol significantly increased the efficiency of single mutation and also allowed facile large single insertions, deletions/truncations and multiple mutations in a single experiment, an option incompatible with the standard QuikChange (TM). Furthermore the new protocol required significantly less parental DNA which facilitated the DpnI digestion after the PCR amplification and enhanced the overall efficiency and reliability. Using our protocol, we generated single site, multiple single-site mutations and a combined insertion/deletion mutations. The results demonstrated that this new protocol imposed no additional reagent costs (beyond basic QuikChange T) but increased the overall success rates.Publisher PDFPeer reviewe
CrY2H-seq: a massively multiplexed assay for deep-coverage interactome mapping.
Broad-scale protein-protein interaction mapping is a major challenge given the cost, time, and sensitivity constraints of existing technologies. Here, we present a massively multiplexed yeast two-hybrid method, CrY2H-seq, which uses a Cre recombinase interaction reporter to intracellularly fuse the coding sequences of two interacting proteins and next-generation DNA sequencing to identify these interactions en masse. We applied CrY2H-seq to investigate sparsely annotated Arabidopsis thaliana transcription factors interactions. By performing ten independent screens testing a total of 36 million binary interaction combinations, and uncovering a network of 8,577 interactions among 1,453 transcription factors, we demonstrate CrY2H-seq's improved screening capacity, efficiency, and sensitivity over those of existing technologies. The deep-coverage network resource we call AtTFIN-1 recapitulates one-third of previously reported interactions derived from diverse methods, expands the number of known plant transcription factor interactions by three-fold, and reveals previously unknown family-specific interaction module associations with plant reproductive development, root architecture, and circadian coordination
Measurement of the 235U(n,f) cross section relative to the 6Li(n,t) and 10B(n,a) standards from thermal to 170 keV neutron energy range at n_TOF
The 235U(n,f) cross section was measured in a wide energy range at n_TOF relative to 6Li(n,t) and 10B(n,alpha), with high resolution and in a wide energy range, with a setup based on a stack of six samples and six silicon detectors placed in the neutron beam. This allowed us to make a direct comparison of the reaction yields under the same experimental conditions, and taking into account the forward/backward emission asymmetry. A hint of an anomaly in the 10÷30 keV neutron energy range had been previously observed in other experiments, indicating a cross section systematically lower by several percent relative to major evaluations. The present results indicate that the evaluated cross section in the 9÷18 keV neutron energy range is indeed overestimated, both in the recent updates of ENDF/B-VIII.0 and of the IAEA reference data. Furthermore, these new high-resolution data confirm the existence of resonance-like structures in the keV neutron energy region. The new, high accuracy results here reported may lead to a reduction of the uncertainty in the 1÷100 keV neutron energy region. Finally, the present data provide additional confidence on the recently re-evaluated cross section integral between 7.8 and 11 eV.Preprin
Developments in the tools and methodologies of synthetic biology.
Synthetic biology is principally concerned with the rational design and engineering of biologically based parts, devices, or systems. However, biological systems are generally complex and unpredictable, and are therefore, intrinsically difficult to engineer. In order to address these fundamental challenges, synthetic biology is aiming to unify a body of knowledge from several foundational scientific fields, within the context of a set of engineering principles. This shift in perspective is enabling synthetic biologists to address complexity, such that robust biological systems can be designed, assembled, and tested as part of a biological design cycle. The design cycle takes a forward-design approach in which a biological system is specified, modeled, analyzed, assembled, and its functionality tested. At each stage of the design cycle, an expanding repertoire of tools is being developed. In this review, we highlight several of these tools in terms of their applications and benefits to the synthetic biology community
Experimental reconstruction of primary hot isotopes and characteristic properties of the fragmenting source in the heavy ion reactions near the Fermi energy
The characteristic properties of the hot nuclear matter existing at the time
of fragment formation in the multifragmentation events produced in the reaction
Zn + Sn at 40 MeV/nucleon are studied. A kinematical focusing
method is employed to determine the multiplicities of evaporated light
particles, associated with isotopically identified detected fragments. From
these data the primary isotopic yield distributions are reconstructed using a
Monte Carlo method. The reconstructed yield distributions are in good agreement
with the primary isotope distributions obtained from AMD transport model
simulations. Utilizing the reconstructed yields, power distribution, Landau
free energy, characteristic properties of the emitting source are examined. The
primary mass distributions exhibit a power law distribution with the critical
exponent, , for isotopes, but significantly deviates from
that for the lighter isotopes. Landau free energy plots show no strong
signature of the first order phase transition. Based on the Modified Fisher
Model, the ratios of the Coulomb and symmetry energy coefficients relative to
the temperature, and , are extracted as a function of A.
The extracted values are compared with results of the AMD
simulations using Gogny interactions with different density dependencies of the
symmetry energy term. The calculated values show a close relation
to the symmetry energy at the density at the time of the fragment formation.
From this relation the density of the fragmenting source is determined to be
. Using this density, the symmetry energy
coefficient and the temperature of fragmenting source are determined in a
self-consistent manner as and
MeV
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A novel NGS library preparation method to characterize native termini of fragmented DNA.
Biological and chemical DNA fragmentation generates DNA molecules with a variety of termini, including blunt ends and single-stranded overhangs. We have developed a Next Generation Sequencing (NGS) assay, XACTLY, to interrogate the termini of fragmented DNA, information traditionally lost in standard NGS library preparation methods. Here we describe the XACTLY method, showcase its sensitivity and specificity, and demonstrate its utility in in vitro experiments. The XACTLY assay is able to report relative abundances of all lengths and types (5' and 3') of single-stranded overhangs, if present, on each DNA fragment with an overall accuracy between 80-90%. In addition, XACTLY retains the sequence of each native DNA molecule after fragmentation and can capture the genomic landscape of cleavage events at single nucleotide resolution. The XACTLY assay can be applied as a novel research and discovery tool for fragmentation analyses and in cell-free DNA
Faddeev treatment of long-range correlations and the one-hole spectral function of O16
The Faddeev technique is employed to study the influence of both
particle-particle and particle-hole phonons on the one-hole spectral function
of O16.
Collective excitations are accounted for at a random phase approximation
level and subsequently summed to all orders by the Faddeev equations to obtain
the nucleon self-energy. An iterative procedure is applied to investigate the
effects of the self-consistent inclusion of the fragmentation in the
determination of the phonons and the corresponding self-energy. The present
results indicate that the characteristics of hole fragmentation are related to
the low-lying states of O16.Comment: 10 pages, 6 figures, 3 tables. Submitted to Phys.Rev.
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