In vivo biosensors: mechanisms,development, and applications

In vivo biosensors can recognize and respond to specific cellular stimuli. In recent years, biosensors have been increasingly used in metabolic engineering and synthetic biology, because they can be implemented in synthetic circuits to control the expression of reporter genes in response to specific cellular stimuli, such as a certain metabolite or a change in pH. There are many types of natural sensing devices, which can be generally divided into two main categories: protein-based and nucleic acid-based. Both can be obtained either by directly mining from natural genetic components or by engineering the existing genetic components for novel specificity or improved characteristics. A wide range of new technologies have enabled rapid engineering and discovery of new biosensors, which are paving the way for a new era of biotechnological progress. Here, we review recent advances in the design, optimization, and applications of in vivo biosensors in the field of metabolic engineering and synthetic biology.     

Founding Pioneers of IVF: Independent innovative researchers generating livebirths within 4 years of the first birth

In this 40th anniversary year of the first IVF live birth, it is pertinent to look at all those teams endeavouring to generate live births from this unique technology and who succeeded within 4 years of the first. There were 9 teams who achieved this and a further 3 who were successful soon after, by the end of 1982. This historical review is compiled by 2 authors who were actively engaged in the field of IVF at the time of the first birth and who have remained active in Reproductive Medicine throughout their professional lives. They bring intimate and relevant knowledge of those pioneer researchers from the early years who can be classified as the "Founding Pioneers" of IVF.     

5个香菇菌株比较研究

研究了５个香菇菌：ＬＤ１ＬＤ７４０５ ＬＤ７９１２：ＬＤ５,ＬＤ６对其栽培性及主要营养成分等重要特性进行了试验比较,从而筛选出ＬＤ６为最优菌株,其现蕾速度快,生产周期短,增重率高等特点,很适宜在北方推广应用。     

A Note on Interval Estimation of the Simple Difference in Data with Correlated Matched Pairs

When we employ cluster sampling to collect data with matched pairs, the assumption of independence between all matched pairs is not likely true. This paper notes that applying interval estimators, that do not account for the intraclass correlation between matched pairs, to estimate the simple difference between two proportions of response can be quite misleading, especially when both the number of matched pairs per cluster and the intraclass correlation between matched pairs within clusters are large. This paper develops two asymptotic interval estimators of the simple difference, that accommodate the data of cluster sampling with correlated matched pairs. This paper further applies Monte Carlo simulation to compare the finite sample performance of these estimators and demonstrates that the interval estimator, derived from a quadratic equation proposed here, can actually perform quite well in a variety of situations.     

Confidence Intervals of the Simple Difference between the Proportions of a Primary Infection and a Secondary Infection,Given the Primary Infection

This paper discusses interval estimation of the simple difference (SD) between the proportions of the primary infection and the secondary infection, given the primary infection, by developing three asymptotic interval estimators using Wald's test statistic, the likelihood‐ratio test, and the basic principle of Fieller's theorem. This paper further evaluates and compares the performance of these interval estimators with respect to the coverage probability and the expected length of the resulting confidence intervals. This paper finds that the asymptotic confidence interval using the likelihood ratio test consistently performs well in all situations considered here. When the underlying SD is within 0.10 and the total number of subjects is not large (say, 50), this paper further finds that the interval estimators using Fieller's theorem would be preferable to the estimator using the Wald's test statistic if the primary infection probability were moderate (say, 0.30), but the latter is preferable to the former if this probability were large (say, 0.80). When the total number of subjects is large (say, ≥200), all the three interval estimators perform well in almost all situations considered in this paper. In these cases, for simplicity, we may apply either of the two interval estimators using Wald's test statistic or Fieller's theorem without losing much accuracy and efficiency as compared with the interval estimator using the asymptotic likelihood ratio test.     

A Note on the Log‐Rank Test in Life Table Analysis with Correlated Observations

Survival data consisting of independent sets of correlated failure times may arise in many situations. For example, we may take repeated observations of the failure time of interest from each patient or observations of the failure time on siblings, or consider the failure times on littermates in toxicological experiments. Because the failure times taken on the same patient or related family members or from the same litter are likely correlated, use of the classical log‐rank test in these situations can be quite misleading with respect to type I error. To avoid this concern, this paper develops two closed‐form asymptotic summary tests, that account for the intraclass correlation between the failure times within patients or units. In fact, one of these two test includes the classical log‐rank test as a special case when the intraclass correlation equals 0. Furthermore, to evaluate the finite‐sample performance of the two tests developed here, this paper applies Monte Carlo simulation and notes that they can actually perform quite well in a variety of situations considered here.     

Cortisol stress response and histopathological alteration index in Clarias gariepinus exposed to sublethal concentrations of Qua Iboe crude oil and rig wash

The histological effect on and stress response of post juvenile Clarias gariepinus exposed to Qua Iboe crude oil and rig wash were investigated. Fish weighing 60–90 g and measuring 16–18 cm were exposed for 7–28 days to 8.00 ml^?1 Qua Iboe crude oil and 0.0018 ml^–1 rig wash, both being 0.1 of the 96 hr LC₅₀. Blood samples of C. gariepinus were collected every seven days and evaluated for stress by measuring cortisol concentration. The gills and liver were studied and scored for Gill Alteration Index (GAI) and Hepatic Alteration Index (HAI), respectively. There was an increase in cortisol level up to the 7th and 14th day among the group exposed to Qua Iboe crude oil, with a decrease on the 21st and 28th day. The rig wash group increased in cortisol level up to the 7th day and decreased slightly on the 14th day, after which the trend became irregular. The toxic effects of the Qua Iboe crude oil and rig wash were time dependent, as shown by the histopathological alteration index (HAI) of gill and liver. After 28 days of exposure, the gills had irreparable damage due to high frequency of cellular necrosis and degeneration, whereas the liver had from moderate to severe damage due to the high frequency of cellular degeneration and inflammation. Qua Iboe crude oil and rig wash are both toxic to C. gariepinus, therefore their indiscriminate discharge to the environment must be discouraged.     

HOP/OB1/NECC1 promoter DNA is frequently hypermethylated and involved in tumorigenic ability in esophageal squamous cell carcinoma

Promoter DNA hypermethylation with gene silencing is a common feature of human cancer, and cancer-prone methylation is believed to be a landmark of tumor suppressor genes (TSG). Identification of novel methylated genes would not only aid in the development of tumor markers but also elucidate the biological behavior of human cancers. We identified several epigenetically silenced candidate TSGs by pharmacologic unmasking of esophageal squamous cell carcinoma (ESCC) cell lines by demethylating agents (5-aza-2'-deoxycitidine and trichostatin A) combined with ESCC expression profiles using expression microarray. HOP/OB1/NECC1 was identified as an epigenetically silenced candidate TSG and further examined for (a) expression status, (b) methylation status, and (c) functional involvement in cancer cell lines. (a) The HOP gene encodes two putative promoters (promoters A and B) associated with two open reading frames (HOPalpha and HOPbeta, respectively), and HOPalpha and HOPbeta were both down-regulated in ESCC independently. (b) Promoter B harbors dense CpG islands, in which we found dense methylation in a cancer-prone manner (55% in tumor tissues by TaqMan methylation-specific PCR), whereas promoter A does not harbor CpG islands. HOPbeta silencing was associated with DNA methylation of promoter B in nine ESCC cell lines tested, and reactivated by optimal conditions of demethylating agents, whereas HOPalpha silencing was not reactivated by such treatments. Forced expression of HOP suppressed tumorigenesis in soft agar in four different squamous cell carcinoma cell lines. More convincingly, RNA interference knockdown of HOP in TE2 cells showed drastic restoration of the oncogenic phenotype. In conclusion, HOP is a putative TSG that harbors tumor inhibitory activity, and we for the first time showed that the final shutdown process of HOP expression is linked to promoter DNA hypermethylation under the double control of the discrete promoter regions in cancer.     

Directed evolution of aniline dioxygenase for enhanced bioremediation of aromatic amines

The objective of this study was to enhance the activity of aniline dioxygenase (AtdA), a multi-component Rieske non-heme iron dioxygenase enzyme isolated from Acinetobacter sp. strain YAA, so as to create an enhanced biocatalyst for the bioremediation of aromatic amines. Previously, the mutation V205A was found to widen the substrate specificity of AtdA to accept 2-isopropylaniline (2IPA) for which the wild-type enzyme has no activity (Ang EL, Obbard JP, Zhao HM, FEBS J, 274:928–939, 2007). Using mutant V205A as the parent and applying one round of saturation mutagenesis followed by a round of random mutagenesis, the activity of the final mutant, 3-R21, was increased by 8.9-, 98.0-, and 2.0-fold for aniline, 2,4-dimethylaniline (24DMA), and 2-isopropylaniline (2IPA), respectively, over the mutant V205A. In particular, the activity of the mutant 3-R21 for 24DMA, which is a carcinogenic aromatic amine pollutant, was increased by 3.5-fold over the wild-type AtdA, while the AN activity was restored to the wild-type level, thus yielding a mutant aniline dioxygenase with enhanced activity and capable of hydroxylating a wider range of aromatic amines than the wild type. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Purple acid phosphatase-like sequences in prokaryotic genomes and the characterization of an atypical purple alkaline phosphatase from Burkholderia cenocepacia J2315

Purple acid phosphatases (PAP) are a group of dimetallic phosphohydrolase first identified in eukaryotes. Bioinformatics analysis revealed 57 prokaryotic PAP-like sequences in the genomes of 43 bacteria and 4 cyanobacteria species. A putative PAP gene (BcPAP) from the bacteria Burkholderia cenocepacia J2315 was chosen for further studies. Synteny analysis showed that this gene is present as an independent gene in most of the members of the genus Burkholderia. The predicted 561 a.a. polypeptide of BcPAP was found to harbour all the conserved motifs of the eukaryotic PAPs and an N-terminal twin-arginine translocation signal. Expression and biochemical characterization of BcPAP in Escherichia coli revealed that this enzyme has a relatively narrow substrate spectrum, preferably towards phosphotyrosine, phosphoserine and phosphoenolpyruvate. Interestingly, this enzyme was found to have a pH optimum at 8.5, rather than an acidic optima exhibited by eukaryotic PAPs. BcPAP contains a dimetallic ion centre composed of Fe and Zn, and site-directed mutagenesis confirmed that BcPAP utilizes the invariant residues for metal-ligation and catalysis. The enzyme is secreted by the wild type bacteria and its expression is regulated by the availability of orthophosphate. Our findings suggest that not all members in the PAP family have acidic pH optimum and broad substrate specificity.