Combined Review: Experiencing the New Genetics: Family and Kinship on the Medical Frontier and Born and Bred: Idioms and New Reproductive Technologies in England

Experiencing the New Genetics: Family and Kinship on the Medical Frontier. Kaja Finkler. Philadelphia: University of Pennsylvania Press, 2000. 296 pp.
Born and Bred: Idioms and New Reproductive Technologies in England. Jeanette Edwards. New York: Oxford University Press, 2000. 264 pp.     

Gene and domain duplication in the chordate Otx gene family: insights from amphioxus Otx

We report the genomic organization and deduced protein sequence of a cephalochordate member of the Otx homeobox gene family (AmphiOtx) and show its probable single-copy state in the genome. We also present molecular phylogenetic analysis indicating that there was single ancestral Otx gene in the first chordates which was duplicated in the vertebrate lineage after it had split from the lineage leading to the cephalochordates. Duplication of a C-terminal protein domain has occurred specifically in the vertebrate lineage, strengthening the case for a single Otx gene in an ancestral chordate whose gene structure has been retained in an extant cephalochordate. Comparative analysis of protein sequences and published gene expression patterns suggest that the ancestral chordate Otx gene had roles in patterning the anterior mesendoderm and central nervous system. These roles were elaborated following Otx gene duplication in vertebrates, accompanied by regulatory and structural divergence, particularly of Otx1 descendant genes.      

18S RIBOSOMAL DNA SEQUENCES INDICATE A MONOPHYLETIC ORIGIN OF CHAROPHYCEAE1

Nuclear-encoded small-subunit (18S) ribosomal RNA genes of Chara australis R. Brown (C. corallina var. nobilis f. nobilis R.D.W.) and Nitella flexilis (L.) Ag. were amplified by polymerase chain reaction, cloned, and completely sequenced. Using structural criteria, the sequences were aligned with 18S ribosomal DNAs (rDNAs) from 11 other chlorophyll b-containing algae and six higher plants (embryophytes). Phylogenetic trees were inferred by distance, neighbor-joining, parsimony, and maximum-likelihood approaches; confidence intervals were estimated by bootstrapping, and nonrandomness of tree structure was confirmed by permutation tests. 18S rDNAs of C. australis and two Nitella species were found to be specifically related and, together with 18S rDNAs of Chlorokybus atmophyticus Geitler, Klebsormidium flaccidum (A. Br.) Silva, Mattox, et Blackwell, and two Coleochaete species, support a robust monophyletic group (Charophyceae). Although most trees favored a specific sister-group relationship between Charophyceae and embryophytes, statistical tests revealed that a sister-group relationship between Charophyceae and Chlorophyceae could not be ruled out. Additional complete sequences from 18S rDNAs of lower land plants may be required to resolve phylogenetic relationships among these organisms.     

Shuttle cloning vectors for the cyanobacterium Anacystis nidulans.

Hybrid plasmids capable of acting as shuttle cloning vectors in Escherichia coli and the cyanobacterium Anacystis nidulans R2 were constructed by in vitro ligation. DNA from the small endogenous plasmid of A. nidulans was combined with two E. coli vectors, pBR325 and pDPL13, to create vectors containing either two selectable antibiotic resistance markers or a single marker linked to a flexible multisite polylinker. Nonessential DNA was deleted from the polylinker containing plasmid pPLAN B2 to produce a small shuttle vector carrying part of the polylinker (pCB4). The two polylinker-containing shuttle vectors, pPLAN B2 and pCB4, transform both E. coli and A. nidulans efficiently and provide seven and five unique restriction enzyme sites, respectively, for the insertion of a variety of DNA fragments. The hybrid plasmid derived from pBR325 (pECAN1) also transforms both E. coli and A. nidulans, although at a lower frequency, and contains two unique restriction enzyme sites.     

Membrane transport by guinea pig peritoneal exudate leukocytes: effect of phagocytosis on hexose and amino acid transport

Short term, carrier mediated transport of D-glucose, L-leucine and L-lysine by guinea pig peritoneal macrophages was characterized. Analysis of the amino acid transport demonstrated two-limbed double reciprocal plots suggesting two transport systems for each amino acid. The low concentration limb of the curves established a Km of 0.1 mM for L-leucine and 0.05 mM for L-lysine; Vmax values were 2.0 and 2.85 nmole/mg protein/90 seconds, respectively. Leucine and lysine were shown to be competitive inhibitors of each other. Further competition studies revealed that other amino acids also had affinity for these carriers. Amino acid transport was found to be sensitive to sulfhydryl active compounds. Colchicine treatment of peritoneal macrophages did not inhibit the transport of the amino acids tested. Preloading macrophages with latex beads or heat-killed staphylococci by phagocytosis stimulated 2-deoxy-D-glucose (2-dOG) uptake markedly, but had no measurable effect on amino acid transport. Although total transport of 2-dOG increased in post-phagocytic macrophages, the kinetics of the system were not altered significantly. The Km for both pre- and post-phagocytic transport of 2-dOG was shown to be 1.2 mM and the Vmax was shown to increase from a pre-phagocytic value of 20 nmoles/mg protein/90 seconds to a post-phagocytic 27 nmoles/mg protein/90 seconds. Phagocytosis of heat-killed staphylococci by guinea pig polymorphonuclear leukocytes (PMNs), however, did not cause an augmentation in hexose transport in the cells. The presence of colchicine during phagocytosis did not alter subsequent uptake of amino acids by the macrophages.     

Cytochemical detection of mannose-specific receptors for glycoproteins with horseradish peroxidase as a ligand

Summary Horseradish peroxidase (HRP), a glycoprotein rich in mannose groups, was used as a ligand to detect receptors for glycoproteins in formalinfixed, frozen sections of rat liver. Specific binding of HRP occurred to surface membranes of sinusoidal cells but not to those of parenchymal cells. The binding sites were visualized after the peroxidatic reaction in erythrocytes had been suppressed by methanol-H₂O₂ and phenylhydrazine, the latter reagent also decreasing the nonspecific background adsorption of HRP. Several factors influencing the reaction were studied systematically. The specific binding of HRP to sinusoidal cells was greatly decreased or abolished when tissue blocks were fixed for longer than 1–2 h in a cold 4% formaldehyde solution and the frozen sections subsequently treated for 30 min in cold methanol. The specific binding of HRP increased when the concentration of HRP in the medium was increased from 10 g/ml to 40 g/ml, when the time of incubation with HRP was increased from 1 h to 4 h, or when the temperature of incubation with HRP was increased from 4°C to 22°C, or from 22°C to 37°C. The specific binding of HRP also increased when the pH of the incubation medium was increased from 7.0 to 10.0. Little or no specific binding of HRP was observed in the absence of added Ca⁺⁺. The binding of HRP was suppressed by 10 mM mannose or 0.004% mannan whereas the suppression of the binding reaction by galactose or galactan required 30–40 times higher concentrations.This work was supported by the Morris A. Kaplan Fund