A simple physiologically relevant double-agar-layer method for post-germination treatment of seedlings

Arabidopsis thaliana is frequently grown on semisolid medium in Petri dishes, for various experiments that usually consist of two stages on two distinct growth media. Seedlings are germinated under favorable conditions followed by their transfer to another medium containing the given treatment(s). This often causes secondary effects on seedlings due to root shock, or direct and unavoidable contact of the shoot with the second medium. We have developed a simple and efficient method for the transfer of seedlings grown on semisolid medium with minimal damage. In this double-agar-layer method, seeds are germinated on a thin growth-medium-containing agar layer. Subsequently, medium blocks containing the embedded seedlings are excised and placed on the second semisolid medium supplemented with the treatment agent. Differential agar concentrations allow easy penetration of the roots into the second medium, but do not allow the shoots to come into contact with it. This unique method offers several advantages over others that are in common use, in which the seedlings are individually transferred to the second medium or alternatively grown on transfer-carrier matrices, such as filter paper, mesh and cellophane. In the presented method, the entire root system faces the growth medium, the shoots are surrounded by air at all growth stages and transfer of the seedlings is much easier. In addition, a large number of seedlings can be transferred in a single step, without stressing the plants or damaging the delicate root system. This method can also be applied to other plant species grown on semisolid media.     

Comparative effects of caffeine on radiation- and heat-induced alterations in cell cycle progression

The effects of 3 mM caffeine on cell cycle progression of HeLa S3 cells exponentially and asynchronously growing in suspension culture were studied following exposure to 6.8 Gy gamma irradiation or 30 min at 45 degrees C hyperthermia. The stathmokinetic method, in which cells are grown in the presence of colcemid for the duration of experiment, in combination with two flow cytometric techniques, propidium iodide staining of DNA and acridine orange staining following acid denaturation of chromatin, were used to determine the fraction of cells in four cell cycle compartments, G1, S, G2, and M. Radiation and caffeine acted in a complementary manner, in which radiation reduced the caffeine-induced delays in cell cycle progression and caffeine prevented completely the radiation-induced accumulation of cells in G2 and mitotic delay. Heat and caffeine had additive effects on alterations in cell cycle progression. Cells containing spontaneous prematurely condensed chromatin were observed transiently immediately following heat exposure. These cells appeared to be in G2 and late S phase.     

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).     

A note on an in vitro test system to compare the bactericidal properties of wound dressings

H olland , K.T. & D avis , W. 1985. A note on an in vitro test system to compare the bactericidal properties of wound dressing. Journal of Applied Bacteriology 59 , 61–63.
An in vitro method of comparing the antimicrobial properties of dressings used on human skin is described and has been tested with two samples of tulle gras. The method has advantages over agar diffusion methods by providing flexibility of test conditions and kinetic data for analysis.     

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.     

HomeoDB2: functional expansion of a comparative homeobox gene database for evolutionary developmental biology

Homeobox gene database (HomeoDB), a manually curated database of homeobox genes and their classification, has been well received since its release in 2008. Here, we report HomeoDB2, an expansion and improvement of the original database that provides greater functionality for the user. HomeoDB2 includes all homeobox loci from 10 animal genomes (human, mouse, chicken, frog, zebrafish, amphioxus, nematode, fruitfly, beetle, honeybee) plus tools for downloading sequences, comparing between species and BLAST searching. HomeoDB2 provides a resource for studying the dynamics of homeobox gene evolution, and is freely accessible at http://homeodb.zoo.ox.ac.uk     

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.