Carbonic anhydrase activity in the blood of adult sheep, fetal sheep and lambs

The activity of carbonic anhydrase (E.C.4.2.1.1) (CA) has been measured in the blood of adult and fetal sheep and lambs. The mean activity in adult sheep was 0.89 enzyme units (EU) per 100 micrograms of Hb. The activity in fetal sheep aged 90 days was just below 20% of this and in fetuses near full term was just under 40% of the mean adult level. The regression line gave an increase of CA activity (per 100 micrograms Hb) of 0.004 EU/day. The appearance of CA in fetal blood normally occurred before any detectable production of adult Hb. One aberrant fetus showed early development of the adult pattern in the red cells, having adult type Hb and adult levels of CA during the period of 116-128 days of fetal age. In the period after birth the CA level in the blood rose rapidly, reaching the adult level 30 days after birth. During this period activity per 100 micrograms HB increased by 0.014 EU/day, significantly faster than during fetal life.     

Virus protein changes and RNA termini alterations evolving during persistent infection

Cloned infectious vesicular stomatitis virus isolated following 5 years of persistent infection of BHK21 cells in vitro exhibits a number of peptide map changes in the G protein (spike glycoprotein), the M protein (membrane matrix protein) and the N protein (nucleocapsid structural protein). Only slight alterations have occurred in the peptide maps of the two VSV polymerase-associated proteins L and NS. Dideoxy sequencing of the 3′ ends of the cloned virus originally used to establish the persistent infection, and of the cloned virus recovered following 5 years of persistence, shows one base substitution in the three base junction between the 3′ leader sequence and the N protein-coding region. Repeated lytic passages of virus recovered from persistent infection led to no oligonucleotide map changes after 30 passages, but two map changes were present after 102 and remained after 133 lytic passages in BHK21 cells in vitro. Only one of these represented reversion to the original map position, and this “mutant” virus still exhibited a temperature-sensitive small plaque phenotype. Finally, the mutated virus recovered after more than $\text{5}\text{1}\text{2}$ years of persistent infection is now so slow-growing that it can establish persistent infection of BHK21 cells in the absence of DI particles (although DI particles are present constantly once the cells recover from the initial cytopathology).     

Selecting representative model micro-organisms

Background  

Micro-biological research relies on the use of model organisms that act as representatives of their species or subspecies, these are frequently well-characterized laboratory strains. However, it has often become apparent that the model strain initially chosen does not represent important features of the species. For micro-organisms, the diversity of their genomes is such that even the best possible choice of initial strain for sequencing may not assure that the genome obtained adequately represents the species. To acquire information about a species' genome as efficiently as possible, we require a method to choose strains for analysis on the basis of how well they represent the species.     

Low-density lipoprotein preparation by combined diafiltration and ultracentrifugation

A method for isolating low-density lipoprotein by combining diafiltration and ultracentrifugation is described. Diafiltration separates plasma components by use of an ultrafiltration membrane that excludes particles of molecular weight greater than 300,000. The retentate is concentrated three- to fourfold by ultrafiltration, allowing large-scale preparation of low-density lipoprotein. Low-density lipoprotein prepared in this manner is similar in physical, chemical, and biologic properties to low-density lipoprotein isolated by sequential density ultracentrifugation alone. When low-density lipoprotein, prepared by either method, was added to human umbilical vein endothelial cell cultures, no cytotoxicity was observed. The techniques described reduce the demand on multiple rotors and ultracentrifuges for large-scale preparation of low-density lipoprotein suitable and often needed for tissue culture studies.     

In vivo expression of mRNA for the Ca++-binding protein SPARC (osteonectin) revealed by in situ hybridization

In situ hybridization is used to survey the tissue-specific and developmental expression of the cloned mouse gene Sparc, coding for a protein homologous to the bovine Ca++-binding protein, osteonectin. High levels of SPARC RNA are found in osteoblasts and odontoblasts. In addition, high grain counts are associated with a variety of other cell types in the embryo and newborn mouse, including parietal endoderm, deciduum, whisker follicles (connective tissue sheath), peripheral nerve trunk, skin (dermis), and stomach (submucosa). Spatially restricted but high levels of SPARC mRNA are also seen in the adult adrenal glands, testis, and ovary. This pattern of differential gene expression demands a reassessment of the function originally proposed for osteonectin, and predicts a much wider role for the protein in a variety of biological processes.