Sequence of the 3''-noncoding and adjacent coding regions of human gamma-globin mRNA.

In cloning human fetal globin cDNA in bacterial plasmids, we obtained a recombinant which contained a fragment of gammg-globin cDNA corresponding to the region from amino acid 99 to the poly A. We determined a sequence of 169 nucleotides which included the complete 3' non-coding region of the gamma-globin mRNA. The codon for amino acid 136 was GCA, indicating that this cloned fragment was derived from the Agamma-globin gene. In conjunction with the surrounding sequences, the GCA codon provides the Agamma-species with a unique CTGCAG hexanucleotide that is recognized by the restriction enzyme Pst I. The 3'-untranslated region of the gamma-globin mRNA consists of 90 nucleotides, and shares little homology with that of the human beta-globin mRNA. As in other mammalian mRNAs, a symmetrical sequence and the hexanucleotide AAUAAA are present.     

The structure, catalytic mechanism and regulation of adenylyl cyclase

The recent structure determinations of the mammalian effector enzyme adenylyl cyclase reveal the structure of its catalytic core, provide new insights into its catalytic mechanism and suggest how diverse signaling molecules regulate its activity.     

A computational study of dsDNA pairs and vibrational resonance in separating water

This article investigates the relationship between molecular sequence and dependent interacting behavior of molecular segment pairs and secondly, sequence dependent, vibrational resonance in surrounding normal saline, protein-free water. The development of a molecular model to explore these systems phenomena, the results of several nanoscale molecular dynamics simulations, and analysis of behavior of interacting ΦX174 double-stranded DNA segment pair models in various configurations are presented. Fourier analysis revealed intriguing vibration frequencies within the solvent plane between the segments, while subsequent frequency domain transformation of the time domain waveforms revealed statistically significant resonating harmonic signals in the THz range.

Electronic supplementary material

The online version of this article (doi:10.1007/s11693-014-9157-3) contains supplementary material, which is available to authorized users.     

Mitochondrial DNA structure and colony expansion dynamics of New Zealand fur seals (Arctocephalus forsteri) around Banks Peninsula

New Zealand fur seals are one of many pinniped species that survived the commercial sealing of the eighteenth and nineteenth centuries in dangerously low numbers. After the enforcement of a series of protection measures in the early twentieth century, New Zealand fur seals began to recover from the brink of extinction. We examined the New Zealand fur seal populations of Banks Peninsula, South Island, New Zealand using the mitochondrial DNA control region. We identified a panmictic population structure around Banks Peninsula. The most abundant haplotype in the area showed a slight significant aggregated structure. The Horseshoe Bay colony showed the least number of shared haplotypes with other colonies, suggesting a different origin of re-colonisation of this specific colony. The effective population size of the New Zealand fur seal population at Banks Peninsula was estimated at approximately 2500 individuals. The exponential population growth rate parameter for the area was 35, which corresponds to an expanding population. In general, samples from adjacent colonies shared 4.4 haplotypes while samples collected from colonies separated by between five and eight bays shared 1.9 haplotypes. The genetic data support the spill-over dynamics of colony expansion already suggested for this species. Approximate Bayesian computations analysis suggests re-colonisation of the area from two main clades identified across New Zealand with a most likely admixture coefficient of 0.41 to form the Banks Peninsula population. Approximate Bayesian computations analysis estimated a founder population size of approximately 372 breeding individuals for the area, which then rapidly increased in size with successive waves of external recruitment. The population of fur seals in the area is probably in the late phase of maturity in the colony expansion dynamic.     

Genetic variation in foraging traits among inbred lines of a predatory mite

Response of predators to herbivore-induced plant volatiles can affect the length of time a predator spends in a prey patch and the probability of a predator finding a new prey patch. Variation in response to herbivore-induced plant volatiles may lead to different foraging decisions among individuals, thereby affecting both within-patch dynamics and between-patch dispersal. We found significant phenotypic and additive genetic variation in two behavioral assays of response to herbivore-induced plant volatiles among inbred isofemale lines of the predatory mite, Phytoseiulus persimilis. In wind-tunnel tests to measure patch residence time, adult female predators from certain lines left prey patches sooner than others when a distant source of herbivore-induced plant volatiles was presented; whereas such variation disappeared when no distant volatiles were presented. In a measure of patch location, certain lines were more likely than others to locate a prey-infested leaf disc; again there was no difference when uninfested leaf discs were used. Patch location was negatively correlated with patch residence. That is, lines that were more likely to leave a prey patch in the presence of distant volatiles were also more likely to find an odor source (ie, prey patch) from a distance of 20 cm. These two foraging-related behaviors are heritable. A continuous distribution of both behaviors indicated that several to many loci may be responsible for these behavioral traits. Our line-crossing experiments suggested that maternal influence could be excluded. Substantial phenotypic variation in two other foraging-related traits, consumption and oviposition, were also detected among inbred lines. Consumption and oviposition were positively correlated; however, the relationship (slope) varied among inbred lines, suggesting that predatory mites vary in food conversion efficiency. A relationship was detected between patch residence and consumption. Patch location, as one important foraging trait, appeared to be negatively related to consumption, suggesting a trade-off between searching for patches and reproduction.     

Fluorescent multiplexing of 3D spheroids: Analysis of biomarkers using automated immunohistochemistry staining platform and multispectral imaging

In preclinical cancer studies, three-dimensional (3D) cell spheroids and aggregates are preferred over monolayer cell cultures due to their architectural and functional similarity to solid tumors. We performed a proof-of-concept study to generate physiologically relevant and predictive preclinical models using non–small cell lung adenocarcinoma, and colon and colorectal adenocarcinoma cell line-derived 3D spheroids and aggregates. Distinct panels were designed to determine the expression profiles of frequently studied biomarkers of the two cancer subtypes. The lung adenocarcinoma panel included ALK, EGFR, TTF-1, and CK7 biomarkers, and the colon and colorectal adenocarcinoma panel included BRAF V600E, MSH2, MSH6, and CK20. Recent advances in immunofluorescence (IF) multiplexing and imaging technology enable simultaneous detection and quantification of multiple biomarkers on a single slide. In this study, we performed IF staining of multiple biomarkers per section on formalin-fixed paraffin-embedded 3D spheroids and aggregates. We optimized protocol parameters for automated IF and demonstrated staining concordance with automated chromogenic immunohistochemistry performed with validated protocols. Next, post-acquisition spectral unmixing of the captured fluorescent signals were utilized to delineate four differently stained biomarkers within a single multiplex IF image, followed by automated quantification of the expressed markers. This workflow has the potential to be adapted to preclinical high-throughput screening and drug efficacy studies utilizing 3D spheroids from cancer cell lines and patient-derived organoids. The process allows for cost, time, and resource savings through concurrent staining of several biomarkers on a single slide, the ability to study the interactions of multiple expressed proteins within a single region of interest, and enable quantitative assessment of biomarkers in cancer cells.     

Increased megakaryopoiesis in cultures of CD34-enriched cord blood cells maintained at 39 degrees C

Based on previous evidence suggesting positive effects of fever on in vivo hematopoiesis, we tested the effect of hyperthermia on megakaryopoiesis (MK) in ex vivo cultures of CD34-enriched cord blood (CB) cells. The cells were cultured at 37 degrees C or 39 degrees C for 14 days in cytokine conditions optimized for megakaryocyte development and analyzed periodically. Compared to 37 degrees C, the cultures maintained at 39 degrees C produced significantly more (up to 10-fold) total cells, myeloid and MK progenitors, and total MKs, and showed accelerated and enhanced MK maturation with increased yields of proplatelets and platelets. This observation could facilitate clinical applications requiring ex vivo expansion of hematopoietic cells.     

An Augmented SMS Intervention to Improve Access to Antenatal CD4 Testing and ART Initiation in HIV-Infected Pregnant Women: A Cluster Randomized Trial

BackgroundLess than one-third of HIV-infected pregnant women eligible for combination antiretroviral therapy (ART) globally initiate treatment prior to delivery, with lack of access to timely CD₄ results being a principal barrier. We evaluated the effectiveness of an SMS-based intervention to improve access to timely antenatal ART.MethodsWe conducted a stepped-wedge cluster randomized trial of a low-cost programmatic intervention in 20 antenatal clinics in Gaborone, Botswana. From July 2011-April 2012, 2 clinics were randomly selected every 4 weeks to receive an ongoing clinic-based educational intervention to improve CD₄ collection and to receive CD₄ results via an automated SMS platform with active patient tracing. CD₄ testing before 26 weeks gestation and ART initiation before 30 weeks gestation were assessed.ResultsThree-hundred-sixty-six ART-naïve women were included, 189 registering for antenatal care under Intervention and 177 under Usual Care periods. Of CD₄-eligible women, 100 (59.2%) women under Intervention and 79 (50.6%) women under Usual Care completed CD₄ phlebotomy before 26 weeks gestation, adjusted odds ratio (aOR, adjusted for time that a clinic initiated Intervention) 0.87 (95% confidence interval [CI]0.47–1.63, P = 0.67). The SMS-based platform reduced time to clinic receipt of CD₄ test result from median of 16 to 6 days (P<0.001), was appreciated by clinic staff, and was associated with reduced operational cost. However, rates of ART initiation remained low, with 56 (36.4%) women registering under Intervention versus 37 (24.2%) women under Usual Care initiating ART prior to 30 weeks gestation, aOR 1.06 (95%CI 0.53–2.13, P = 0.87).ConclusionsThe augmented SMS-based intervention delivered CD₄ results more rapidly and efficiently, and this type of SMS-based results delivery platform may be useful for a variety of tests and settings. However, the intervention did not appear to improve access to timely antenatal CD₄ testing or ART initiation, as obstacles other than CD₄ impeded ART initiation during pregnancy.