Agar plates

Thermoduric, Thermophilic, andPsychrophihc Bacteria. Thermoduric bacteria survive but do not grow at pasteurization temperatures. They are largely non spore-forming, heat-resistant types that develop on surfaces of unclean equipment. These bacteria are determined by subjecting a sample to laboratory pasteurization and examining it by the agar plate method. 

Thermophilic bacteria are able to grow at 55°C. They are spore-forming bacilli that can enter milk from a variety of farm sources. Tbermophiles grow ia milk held at elevated temperatures. Their presence ia milk is determined by means of the agar plate method and iacubation at 55°C. 

Add 10 g soil to 100ml liquid Medium A. Incubate with shaking in darkness for 24h at 22°C. Spread 10 x 0.1 ml volumes on agar plates of Medium A and incubate at 22°C until colonies appear. 

Three 1 litre baffled flasks, each containing 100 ml medium, are inoculated with cells from one agar plate suspended in 10 ml saline and incubated at 30°C on a rotary shaker (for optimum supply of oxygen). This provides sufficient biomass to inoculate the bioreactor. 

Take a loop full of the creamy culture off the agar plate. Dip the loop into a 250 ml flask containing 100ml of 5.0g-l 1 glucose and 1 g-1 1 yeast extract. Swirl the loop in the nutrient solution to dislodge the selected culture from the loop. 

In each cycle, the library of mutated genes is first inserted in a standard bacterial host such as Escherichia coli or Bacillus subtilis. Subsequently, bacterial colonies are plated out on agar plates and harvested individually by a colony picker. Each colony is placed in a separate well of a microtiter plate containing nutrient broth, so that the bacteria grow and produce the protein of interest. Because each colony originates... 

In this technique the concentration of a drug in an agar plate may (theoretically) be varied infinitely between zero and a given maximum. To perform the test, nutrient agar is melted, the solution under test added, and the mixture poured into a sterile Petri dish and allowed to set in the form of a wedge (Fig. 11.6A). 

To test their inhibitory power, the powders may be dusted onto the surfaee of seeded agar plates, using the inert diluent as a control and noting the extent of growth. 

Solutions of the substrate have been prepared, for example, in acetone or diethyl ether, and added to or spread over the surface of agar plates either before or after inoculation (Sylvestre 1980 Shiaris and Cooney 1983). 

A solution of the substrate in ethanol may be mixed with the bacterial suspension in agarose and poured over agar plates of the base medium (Bogardt and Hemmingsen 1992). 

For organisms that grow poorly in liquid medium, it may be difficult to obtain sufficient quantities of cells cultures may then be grown on the surface of agar plates and the cells removed by scraping. 

Due to the inherent variability of these assays either by agar-plate diffusion measurement or turbidimetry measurement, the fiducial hmits are calculated according... 

Figure 3 Root fingerprints of Pseudomimets sp. associated with barley seedlings showing the production of siderophore by actively growing bacteria located in the zone of elongation behind the root tips. Root.s were pressed on to an iron-deficient minimal medium selective for Pseudomonas. After growth of the colonies, the production of siderophore was visualized by exposure of the agar plate to ultraviolet light, which causes the siderophore to Huoresce.

Clearing zones on agar plates Most probable number methods... 

Duplicate counts are made using serial dilutions up to 10-6 and drop plates. Solutions are then spotted onto blood agar plates and incubated at 37 °C for 18 hours after which the number of colony forming units is determined. To pass the BA Challenge Test there must be no growth from the aliquots taken at 15 minutes or more from the 1 400 and 1 1600 dilutions. 

Figure 5.10. Protein complementation assay using murine DHFR. The F[l,2] and F[3] fragments are each fused to the homodimerizing GCN4 leucine zipper protein. A. Transformation of both Z-F[l,2] and Z-F[3] constructs results in reconstituted DHFR and growth of E. coh on agar plates containing trimethoprim. B. Transformation of Z-F[l,2] or Z-F[3] alone does not result in trimethoprim resistant E. coli cells. Figure adapted from Pelletier et al. (1998).

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