An “instant gene bank” method for heterologous gene cloning: complementation of two Aspergillus nidulans mutants with Gaeumannomyces graminis DNA

We present a novel technique for gene cloning by complementation of mutations in Aspergillus nidulans with DNA from a heterologous organism, Gaeumannomyces graminis. This technique bypasses the time-consuming and difficult construction of gene libraries, making it both rapid and simple. The method relies on recombination between a fungal replicating vector pHELP1 and linear G. graminis genomic DNA during co-transformation. We were able to complement two out of seven A. nidulans mutants tested and to rescue transforming DNA from both in Escherichia coli. Complementation of the A. nidulans argB mutation resulted from integration of 8–10 kb segments of G. graminis DNA into pHELP1. The complementation of the A. nidulans pyrG mutation resulted from a complex rearrangement. Complementing DNA was shown to originate from G. graminis, and was capable of retransforming the original mutants to give the expected phenotype.     

‘Initial state’ coordinations reproduce the instant flexibility for human walking

An important feature of human locomotor control is the instant adaptability to unpredictable changes of conditions surrounding the locomotion. Humans, for example, can seamlessly adapt their walking gait following a sudden ankle impairment (e.g., as a result of an injury). In this paper, we propose a theoretical study of the mechanisms underlying flexible locomotor control. We hypothesize that flexibility is achieved by modulating the posture at the beginning of the stance phase—the initial state. Using a walking model, we validate our hypothesis through computer simulations     

An automated method for the analysis of ‘particulate’ carbon and nitrogen in natural waters

A method is described for the measurement of the carbon and nitrogen content of particulate material in natural waters. Particulate material is separated by filtration through GF/C filters. The dried filter is encapsulated in silver foil using a purpose made press. Analysis is carried out using high temperature combustion with thermal conductivity detection of emission gasses. Analytical performance characteristics obtained with both standards and natural materials are given.     

An extension of the ‘marker regression’ method to interactive QTL

A regression mapping approach to localize simultaneously two unlinked interactive QTL is proposed. It extends the simple marker regression method of Kearsey and Hyne (1994) to the case of two linkage groups, each with a single QTL having no additive (main) effect, and acting only through epistasis (interaction between a pair of QTL). It allows the localization of the two QTL on the two linkage groups involved by a least square method, which is theoretically more precise than the classical study of interactions between pairs of markers. An application of the method to interactive QTL controlling plant height in bread wheat is presented. Further possible developments are briefly discussed.     

A modified version of the digestion–ligation cloning method for more efficient molecular cloning

Here we describe a modified version of the digestion–ligation approach for efficient molecular cloning. In comparison with the original method, the modified method has the additional steps of gel purification and a second ligation after the first ligation of the linearized vector and DNA insert. During this process, the efficiency and reproducibility could be significantly improved for both stick-end cloning and blunt-end cloning. As an improvement of the very important molecular cloning technique, this method may find a wide range of applications in bioscience and biotechnology.     

Stochasticity or the fatal ‘imperfection’ of cloning

The concept of clone is analysed with the aim of exploring the limits to which a phenotype can be said to be determined geneticaly. First of all, mutations that result from the replication, topological manipulation or lesion of DNA introduce a source of heritable variation in an otherwise identical genetic background. But more important, stochastic effects in many biological processes may superimpose a phenotypic variation which is not encoded in the genome. The source of stochasticity ranges from the random selection of alleles or whole chromosomes to be expressed in small cell populations, to fluctuations in processes such as gene expression, due to limiting amounts of the players involved. The picture emerging is that the term clone is a statistical over-simplification representing a series of individuals having essentially the same genome but capable of exhibiting wide phenotypic variation. Finally, to what extent fluctuations in biological processes, usually thought of as noise, are in fact signal is also discussed.     

Molecular cloning and expression of an expansin-like gene in ‘Navel’ orange fruit during postharvest stresses

In a search for differentially expressed genes in peel pitting of ‘Navel’ orange fruit (Citrus sinensis L. Osbeck), a cDNA subtraction library was constructed and a sequence encoding expansin-like gene was isolated and identified as pitting related gene. Based on sequence information derived from this fragment, a full-length cDNA (CsEXP, GenBank accession no. FJ769424) of 1,083 nucleotides encoding expansin was isolated from ‘Navel’ orange by RACE approaches. CsEXP encoded a protein of 254 amino acid residues with an open reading frame located in the region between 52 and 816 bp. The calculated molecular weight of the mature protein was 27.05 kDa and theoretical isoelectric point was 7.93. The deduced protein contained conserved domains of expansin: the histidine-phenylalanine-aspartate motif in central portion, cysteine residues in N-terminus, and tryptophan residues in C-terminal region. The expression of CsEXP was higher in pitting than the control. Exposure of fruit to stresses, including wounding, anoxia, low temperature (4°C), and treatment with ethylene, increased CsEXP mRNA levels in comparison with the control untreated fruit, whereas high temperature (40°C) decreased its mRNA levels. Since low temperature, low oxygen and wounding were suspected factors inducing peel pitting of citrus fruit. The present results provided us a clue that CsEXP may play a role in response to peel pitting related stresses.     

Integrative factor analysis — An unsupervised method for quantifying cross-study consistency of gene expression data

Integrative analyses of multiple gene expression studies are frequently performed. In the setting of two studies, integrative correlation (IGC) can be used to assess the consistency of co-expression of a given gene. For three or more studies, an extension of IGC gives a global score per gene. We propose to extend IGC and use factor analysis to assess the study-specific consistency of co-expression of genes when there are three or more studies, possibly on different platforms. Our method is able to identify studies whose expression patterns are different from others. Filtering genes based on our score is shown to improve the concordance of association with phenotype across studies.     

Behaviour of a ‘sticky-desynaptic’ mutant in pearl millet

In the progeny after selfing of a normally open pollinated variety (L.S. 326-3) of pearl millet (Pennisetum americanum (L.) Leeke, n=7) one plant exhibited desynapsis, chromosome stickiness and high sterility. Meiosis was normal until diplotene. Thereafter, it was characterized by dissociation of bivalents into univalents and formation of nonspecific congregations of chromosomes at diakinesis, shrinkage of cytoplasm and occurrence of unoriented sticky chromatin masses at metaphase I, relaxation of stickiness, unbalanced chromosome numbers at the poles and laggards at anaphase I, and presence of other irregularities in subsequent stages. Meiosis was completed. Male and female sterility was high. This meiotic mutant thus has multiple effects and is inherited as a monogenic recessive and designated as st.     

Molecular cloning and sequence analysis of the ribosomal ‘A’ protein gene from the archaebacterium,Halobacterium halobium

The ribosomal ‘A’ protein gene of Halobacterium halobium has been cloned and the nucleotide sequence of the DNA fragment containing the ‘A’ protein gene has been determined. The amino-acid sequence of the protein deduced from the nucleotide sequence was established from manual sequence analysis of the protein and structural data provided by peptides derived from cleavage of the protein with various proteinases. The ‘A’ protein consisted of 114 amino acids with a molecular weight of 11562 and was characterized mainly by a high amounts of alanine and acidic amino acid in the C-terminal half of the molecule. The coding sequence of the gene was preceded by a predicted Shine-Dalgarno sequence and two terminal codons. There was no intron or insertion sequence in the coding sequence. Following the terminal codon of the ‘A’ gene, there was a structure reminiscent of the Escherichia coli rho-independent terminator. The G + C content of the coding sequence was found to be 71%. Inspection of the codon usage for the ‘A’ gene revealed 85% preference for G or C at the third codon position.     

Special Issue on‘‘Biomarkers for Diseases’’

<正>The journal Genomics ProteomicsBioinformatics(GPB)is now inviting submissions for a special issue(to be published in the winter of 2015)on the topic of‘‘Biomarkers for Diseases’’.As an emerging index,disease biomarkers have demonstrated the potential application in diagnosis and prognosis.The detection of the disease indicators at molecular level,DNA,RNA     

λN gene expression regulated by translation termination in ribosome L24 mutant

Although the initiation and elongation steps of protein synthesis have been extensively char-acterized in Escherichia coli (E. coli), the translation termination is still less well understood. However, recent experiment result might have provided some hints for our deeper understanding of the termination mechanism. (i) Not only does the translation stop codon act as a signal for ter-mination, but also its context influences the translation termination[13]; (ii) the structure similar-ity betwee…