Meiotic Behavior and Linkage Relationships in the Secondarily Homothallic Fungus Agaricus Bisporus

This study followed the transmission of 64 segregating genetic markers to 52 haploid offspring, obtained from both homokaryotic and heterokaryotic meiospores, of a cross (AG 93b) of Agaricus bisporus, the commonly cultivated ``button mushroom.' The electrophoretic karyotypes of the AG 93b component nuclei were determined concurrently (n = 13). Eleven distinct linkage groups were identified by two-point analysis. DNA-DNA hybridization showed that nine of these corresponded to unique chromosome-sized DNAs. Two other chromosomal DNAs were marked with nonsegregating markers, including the rDNA repeat. Two remaining chromosomes remained unmarked but hybridized to repeated-sequence probes. Cross 93b had an essentially conventional meiosis in which both independent assortment and joint segregation of markers occurred, but in which crossing over was infrequent over much of the mapped genome. The 48 homokaryotic spore-offspring had overall crossover frequencies that were similar to, but possibly slightly less than, those of three homokaryon constituents of heterokaryotic spore-offspring. These data provide support for our earlier cytogenetic model of sporogenesis in A. bisporus, that explains why heterokaryotic spore-offspring usually appear to exhibit no recombination. No evidence favoring an alternative, mitotic model of sporogenesis was found. The resulting genetic map appears to survey the genome extensively and for the first time permits localization of loci determining economically important traits in this fungal crop species. Large differences in the vigor of homokaryotic offspring were correlated with the inheritance of certain chromosome segments and were also often associated with significant departures from Mendelian segregation ratios.     

A novel wound-inducible extensin gene is expressed early in newly isolated protoplasts of Nicotiana sylvestris

A cDNA clone (6PExt 1.2) encoding a novel extensin was isolated from a cDNA library made from 6 h old mesophyll protoplasts of Nicotiana sylvestris. The screening was performed with a heterologous probe from carrot. The encoded polypeptide showed features characteristic of hydroxyproline-rich glycoproteins such as Ser-(Pro)₄ repeats and a high content in Tyr and Lys residues. The presence of four Tyr-X-Tyr-Lys motifs suggests the possibility for intramolecular isodityrosine cross-links whereas three Val-Tyr-Lys motifs may participate in intermolecular cross-links. The analysis of genomic DNA gel blots using both the N. sylvestris and the carrot clones as probes showed that the 6PExt 1.2 gene belongs to a complex multigene family encoding extensin and extensin-related polypeptides in N. sylvestris as well as in related Nicotianeae including a laboratory hybrid. This was confirmed by the analysis of RNA gel blots: a set of mRNAs ranging in size from 0.3 kb to 3.5 kb was found by the carrot extensin probe. The 6PExt 1.2 probe found a 1.2 kb mRNA in protoplasts and in wounded tissues as well as a 0.9 kb mRNA which seemed to be stem-specific. The gene encoding 6PExt 1.2 was induced by wounding in protoplasts, in leaf strips and after Agrobacterium tumefaciens infection of stems.     

Empirical choice of sampling procedures for optimal research design in the longitudinal study of primate behavior

A method is suggested to evaluate on an empirical basis sampling plans for the longitudinal study of primate behavior in those very common situations in which the mathematical structure of behavior is unknown. The method is based on a randomization procedure applied to a pilot sample of the behavior organized so that cost of implementation of a sampling plan can be evaluated vis á vis the sampling error intrinsic to the plan.     

Occurrence of 28- and 27-Carbon ecdysteroids and sterols in developing worker pupae of the leaf-cutting ant acromyrmex octospinosus (reich) (hymenoptera,formicidae: Attini)

The major ecdysteroids in large worker pupae of the leaf-cutting ant Acromyrmex octospinosus were characterized at the peak ecdysteroid concentration by using high-performance liquid chromatography, enzyme immunoassay, and mass spectrometry. In decreasing amounts, they were determined to be makisterone A, an unidentified C₂₈ ecdysteroid bearing a molecular weight of 494, 20-hydroxyecdysone (ratio of 1 to 6 as compared to makisterone A), and putative but negligible ecdysone. The presence of both C₂₈ and C₂₇ ecdysteroids is discussed in relation to the content of 4-desmethylsterols determined by gas chromatography and mass spectrometry to be ergosta-5,7,24 (28)-trien-3β-ol, ergosterol, ergosta-5,7-dien-3β-ol and ergosta-7,24(28)-dien-3β-ol for the main sterols, and with a small amount of cholesterol.     

Mapping of Escherichia coli RNA polymerase binding sites on 2-micrometers DNA from Saccharomyces cerevisiae. Heterogeneity within the inverted duplication and evidence for an eukaryotic invertible DNA sequence.

The 2-μm DNA plasmids from Saccharomyces cerevisiae strain H1 and strain HQ/5C were analyzed by electron microscopy for the presence of Escherichia coli RNA polymerase binding sites. On native 2-μm DNA isolated from strain HQ/5C five RNA polymerase binding sites were detected. One further site was mapped on cloned 2-μm DNA type 23 from S. cerevisiae strain H1. This additional site is located at a distance of 2.15 kilobases from EcoRI site B inside one of the inverted duplication (id) sequences. No such binding site could be detected in the other id sequence of the type 23 molecule, thus indicating that the two id sequences of strain H1 differ in at least one short region. The location of the id sequence carrying the RNA polymerase binding site was analyzed in native 2-μm DNA isolated from strain H1 and found to be present on HindIII fragment 2 and absent from HindIII fragment 5. This indicates that at least a part of the id sequences has a fixed position with respect to the unique S segment and further suggests a site specific recombination mechanism for the inversion of one of the unique segments. As a control for the specificity of RNA polymerase binding, we have mapped binding sites on vectors pBR313 and pBR322. The location of the E. coli RNA polymerase binding sites on 2-μm DNA is discussed in relation to the DNA regions expressed in E. coli minicells.     

Synthesis of high molecular weight polypeptides in Escherichia coli minicells directed by cloned Saccharomyces cerevisiae 2-μm DNA

The minicell-producing Escherichia coli strain P 678-54 was transformed with a series of defined PTY chimeric plasmids consisting of yeast 2-μm DNA and E. coli plasmid pCR1. In minicells the integrated 2-μm DNA from yeast directed specifically the synthesis of six polypeptides with apparent molecular weights of 15 000, 17 500, 20 000, 22 000, 37 000, and 48 000. The specificity of five other polypeptides, which cover a molecular weight range of 19 000 to 28 000, has not yet been established with certainty. Neither the orientation of the integrated DNA, nor the inversion which distinguishes the two structural forms of 2-μm DNA affected the polypeptides synthesized. However, integration at a given EcoRI site appeared to be correlated with the absence of one particular polypeptide band; this suggests that at least one of these sites is located in an expressed region of the DNA.     

Principles in the assembly of annelid erythrocruorins.

Erythrocruorins are giant extracellular respiratory proteins found freely dissolved in the blood of annelids. We present here results from our ultracentrifugation, electron microscopy, spectroscopy, and diffraction experiments on these erythrocruorins. These data are rationalized in terms of a three-dimensional model of the quaternary structure. The proposed structure is arranged in a hierarchy of symmetry. The implications of this structure for the assembly process are considered with special attention to uniqueness and self-limitation. The hypothesis is consistent with observations not used in its construction and it serves as a working hypothesis to focus further experimentation.     

A comparison of major histocompatibility complex SNPs in Han Chinese residing in Taiwan and Caucasians

Summary Genetic dissection of complex diseases is both important and challenging. The human major histocompatibility complex is involved in many human diseases and genetic mechanisms. This highly polymorphic chromosome region has been extensively studied in Caucasians but not as well in Asians. Thus, we compared genotypic distributions, linkage disequilibria and haplotype blocks between Caucasian and Taiwan’s Han Chinese populations. Moreover, we investigated the population admixture and phylogenetic system in Han Chinese residing in Taiwan. The results show that Taiwan’s Han Chinese differ drastically in genotypic information compared with Caucasians but are relatively homogeneous among the three major ethnic subgroups, Minnan, Hakka and Mainlanders. Differences in allele frequency (AF) between Taiwanese and Caucasians in some disease-associated loci may reveal clues to differences in disease prevalence. The results of ethnic heterogeneity imply that public databases should be used with caution in cases where the study population(s) differs from the population characterized in the database. The high homogeneity we observed among the Taiwanese subpopulations mitigates the possibility of spurious association caused by ignoring population stratification in Taiwanese disease gene association studies. These results are useful for understanding our genetic background and designing future disease gene mapping studies.Electronic Supplementary Material Supplementary material is available for this article at and is accessible for authorized users.     

Large-scale recording of neurons by movable silicon probes in behaving rodents

A major challenge in neuroscience is linking behavior to the collective activity of neural assemblies. Understanding of input-output relationships of neurons and circuits requires methods with the spatial selectivity and temporal resolution appropriate for mechanistic analysis of neural ensembles in the behaving animal, i.e. recording of representatively large samples of isolated single neurons. Ensemble monitoring of neuronal activity has progressed remarkably in the past decade in both small and large-brained animals, including human subjects. Multiple-site recording with silicon-based devices are particularly effective because of their scalability, small volume and geometric design. Here, we describe methods for recording multiple single neurons and local field potential in behaving rodents, using commercially available micro-machined silicon probes with custom-made accessory components. There are two basic options for interfacing silicon probes to preamplifiers: printed circuit boards and flexible cables. Probe supplying companies (http://www.neuronexustech.com/; http://www.sbmicrosystems.com/; http://www.acreo.se/) usually provide the bonding service and deliver probes bonded to printed circuit boards or flexible cables. Here, we describe the implantation of a 4-shank, 32-site probe attached to flexible polyimide cable, and mounted on a movable microdrive. Each step of the probe preparation, microdrive construction and surgery is illustrated so that the end user can easily replicate the process.