Transformation and Storage of Competent Yeast Cells

Author(s):  
M. R. Altherr ◽  
L. A. Quinn ◽  
C. I. Kado ◽  
R. L. Rodroguez
Keyword(s):  
2013 ◽  
Vol 76 (12) ◽  
pp. 2024-2030 ◽  
Author(s):  
JESSE M. ZUEHLKE ◽  
CHARLES G. EDWARDS

Brettanomyces is a major threat to red wine quality, causing off-odors such as “medicinal,” “barnyard,” or even “sewage” during aging. Although sulfites (SO2) are used to limit spoilage by these yeast cells, reduced storage temperatures may lessen SO2 requirements. To test this hypothesis, a 4 × 4 factorial experimental design with molecular SO2 (mSO2) concentration (0.0, 0.2, 0.5, or 1.1 mg/liter) and storage temperature (22, 18, 15, or 10°C) was devised. Of three strains evaluated, B5 was the lone strain to regain culturability following exposure to 0.5 mg/liter mSO2 (18°C), whereas only F3 remained culturable in the absence of mSO2 at 10°C. Application of fluorescence microscopy using two different probes and quantitative PCR assays revealed only a 2-log reduction in metabolically active cells from wines with SO2 that were not culturable on nonselective media. Culturability in these wines eventually returned regardless of the concentration of mSO2 present. In addition, 4-ethylphenol production ceased upon addition of SO2. These findings provide additional support that Brettanomyces can enter a “viable-but-not-culturable” state upon exposure to sulfites. Given the diversity among strains, maintaining conditions of ≤15°C and ≥0.4 mg/liter mSO2 will help limit spoilage by Brettanomyces but will not lead to its complete eradication.


Author(s):  
E. Keyhani

The matrix of biological membranes consists of a lipid bilayer into which proteins or protein aggregates are intercalated. Freeze-fracture techni- ques permit these proteins, perhaps in association with lipids, to be visualized in the hydrophobic regions of the membrane. Thus, numerous intramembrane particles (IMP) have been found on the fracture faces of membranes from a wide variety of cells (1-3). A recognized property of IMP is their tendency to form aggregates in response to changes in experi- mental conditions (4,5), perhaps as a result of translational diffusion through the viscous plane of the membrane. The purpose of this communica- tion is to describe the distribution and size of IMP in the plasma membrane of yeast (Candida utilis).Yeast cells (ATCC 8205) were grown in synthetic medium (6), and then harvested after 16 hours of culture, and washed twice in distilled water. Cell pellets were suspended in growth medium supplemented with 30% glycerol and incubated for 30 minutes at 0°C, centrifuged, and prepared for freeze-fracture, as described earlier (2,3).


Author(s):  
R. C. Gonzalez

Interest in digital image processing techniques dates back to the early 1920's, when digitized pictures of world news events were first transmitted by submarine cable between New York and London. Applications of digital image processing concepts, however, did not become widespread until the middle 1960's, when third-generation digital computers began to offer the speed and storage capabilities required for practical implementation of image processing algorithms. Since then, this area has experienced vigorous growth, having been a subject of interdisciplinary research in fields ranging from engineering and computer science to biology, chemistry, and medicine.


Author(s):  
K. J. Böhm ◽  
a. E. Unger

During the last years it was shown that also by means of cryo-ultra-microtomy a good preservation of substructural details of biological material was possible. However the specimen generally was prefixed in these cases with aldehydes.Preparing ultrathin frozen sections of chemically non-prefixed material commonly was linked up to considerable technical and manual expense and the results were not always satisfying. Furthermore, it seems to be impossible to carry out cytochemical investigations by means of treating sections of unfixed biological material with aqueous solutions.We therefore tried to overcome these difficulties by preparing yeast cells (S. cerevisiae) in the following manner:


Author(s):  
E. Keyhani

The mutagenic effect of ethidium bromide on the mitochondrial DNA is well established. Using thin section electron microscopy, it was shown that when yeast cells were grown in the presence of ethidium bromide, besides alterations in the mitochondria, the plasma membrane also showed alterations consisting of 75 to 110 nm-deep pits. Furthermore, ethidium bromide induced an increase in the length and number of endoplasmic reticulum and in the number of intracytoplasmic vesicles.Freeze-fracture, by splitting the hydrophobic region of the membrane, allows the visualization of the surface view of the membrane, and consequently, any alteration induced by ethidium bromide on the membrane can be better examined by this method than by the thin section method.Yeast cells, Candida utilis. were grown in the presence of 35 μM ethidium bromide. Cells were harvested and freeze-fractured according to the procedure previously described.


Author(s):  
John W. Roberts ◽  
E. R. Witkus

The isopod hepatopancreas, as exemplified by Oniscus ascellus. is comprised of four blind-ending diverticula. The regenerative cells at the tip of each diverticula differentiate into either club-shaped B-cells, which serve a secretory function, or into conoid S-cells, which serve in the absorption and storage of nutrients.The glandular B-cells begin producing secretory material with the development of rough endoplasmic reticulum during their process of maturation from the undifferentiated regenerative cells. Cytochemical and morphological data indicate that the hepatopancreas sequentially produces two types of secretory material within the large club-shaped cells. The production of the carbohydrate-like secretory product in immature cells seems to be phased out as the production of the osmiophilic secretion was phased in as the cell matured.


Author(s):  
Hirano T. ◽  
M. Yamaguchi ◽  
M. Hayashi ◽  
Y. Sekiguchi ◽  
A. Tanaka

A plasma polymerization film replica method is a new high resolution replica technique devised by Tanaka et al. in 1978. It has been developed for investigation of the three dimensional ultrastructure in biological or nonbiological specimens with the transmission electron microscope. This method is based on direct observation of the single-stage replica film, which was obtained by directly coating on the specimen surface. A plasma polymerization film was deposited by gaseous hydrocarbon monomer in a glow discharge.The present study further developed the freeze fracture method by means of a plasma polymerization film produces a three dimensional replica of chemically untreated cells and provides a clear evidence of fine structure of the yeast plasma membrane, especially the dynamic aspect of the structure of invagination (Figure 1).


Author(s):  
J. M. Paque ◽  
R. Browning ◽  
P. L. King ◽  
P. Pianetta

Geological samples typically contain many minerals (phases) with multiple element compositions. A complete analytical description should give the number of phases present, the volume occupied by each phase in the bulk sample, the average and range of composition of each phase, and the bulk composition of the sample. A practical approach to providing such a complete description is from quantitative analysis of multi-elemental x-ray images.With the advances in recent years in the speed and storage capabilities of laboratory computers, large quantities of data can be efficiently manipulated. Commercial software and hardware presently available allow simultaneous collection of multiple x-ray images from a sample (up to 16 for the Kevex Delta system). Thus, high resolution x-ray images of the majority of the detectable elements in a sample can be collected. The use of statistical techniques, including principal component analysis (PCA), can provide insight into mineral phase composition and the distribution of minerals within a sample.


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