Chromosomal DNA preparation from yeasts of biotechnological importance (original) (raw)
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A procedure is described for the detection of specific DNA sequences in Saccharomyces cerevisiae. This method allows a rapid screening of a large number of yeast colonies. The yeast cells of each colony, grown on nitrocellulose filters, are converted, in situ, to protoplasts by snail enzyme, and are then lysed and their DNAs are denatured and fixed on the filter. The presence of the specific DNA sequence is detected directly on the filter by hybridization with a radioactive cRNA.
The General Properties of a Strain of Yeast with Less DNA than Normal
Journal of General Microbiology, 1972
A strain of yeast called 2 n~ is described which behaved genetically as a diploid, but had approximately 18 % less DNA than a normal diploid. It produced apparently normal haploid ascospore cultures, which had approximately 20 yo less DNA than normal haploids. The growth of 2 n~ from low yeast concentrations was normal, but at high concentrations it was abnormal; viability was not affected. The timing of DNA synthesis in a synchronously dividing culture was in the usual position in the growth cycle for yeast. DNA was estimated by the diphenylarnine reaction; there was no special effect on this reaction, peculiar to 2 n~ and its ascospore cultures. DNA values were confirmed by Azao measurements. The yeast 2 n~ was more sensitive to both gamma and U.V. radiations than were normal diploids and there was some indication that defective repair processes were involved.
A DNA extraction and purification method for several yeast genera
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Biodiversity and other branches of biological research require rapid and inex- pensive methods to extract and purify DNA, able to process several samples simultaneous- ly, thus allowing an extensive approach to the molecular study of diversity in nature. One problem is the necessity of choosing between rapid methods able to produce relatively crude products or time-consuming procedure yielding nucleic acids
Assignment of cloned genes to the seven electrophoretically separated Candida albicans chromosomes
Molecular and Cellular Biology, 1988
By using orthogonal-field alternating gel electrophoresis (OFAGE), field-inversion gel electrophoresis (FIGE), and contour-clamped homogeneous field gel electrophoresis (CHEF), we have clearly resolved 11 chromosomal bands from various Candida albicans strains. OFAGE resolves the smaller chromosomes better, while FIGE, which under our conditions causes the chromosomes to run in the reverse order of OFAGE, is more effective in separating the larger chromosomes. CHEF separates all chromosomes under some conditions, but these conditions do not often resolve homologs. The strains examined are highly polymorphic for chromosome size. Fourteen cloned Candida genes, isolated on the basis of conferral of new properties to or complementation of auxotrophic deficiencies in Saccharomyces cerevisiae, and three sequences of unknown function have been hybridized to Southern transfers of CHEF, FIGE, and OFAGE gels. Four sets of resolvable bands have been shown to be homologous chromosomes. On the b...
Applied and Environmental Microbiology, 2009
Yeasts used in the production of lagers contain complex allopolyploid genomes, resulting from the fusion of two different yeast species closely related to Saccharomyces cerevisiae and Saccharomyces bayanus. Recombination between the homoeologous chromosomes has generated a number of hybrid chromosomes. These recombination events provide potential for adaptive evolution through the loss or gain of gene function. We have examined the genotypic and phenotypic effects of one of the conserved recombination events that occurred on chromosome XVI in the region of YPR159W and YPR160W. Our analysis shows that the recombination event occurred within the YPR160W gene, which encodes the enzyme glycogen phosphorylase and generates a hybrid gene that does not produce mature mRNA and is nonfunctional due to frameshifts in the coding region. The loss of function of the hybrid gene leads to glycogen levels similar to those found in haploid yeast strains. The implications for the control of glycogen levels in fermentative yeasts are discussed.
Yeast Genetics and Biotechnological Applications
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Contents for conducting classical genetics. Various vector systems available, foreign genes expressed and the limitations as a host will be discussed. Finally, the use of various yeast enzymes in biotechnology sector will be reviewed.
The ploidy determination of the biotechnologically important yeast Candida utilis
Journal of Applied Genetics, 2020
Yeast Candida utilis is considered to be a potentially advantageous expression system for production of recombinant proteins utilizable for industrial and pharmaceutical purposes. As the scientific literature is not consistent in the ploidy of this yeast, in this work, we focused on resolving the problem via several methods such as the copy number determination of maltase gene by multiplex PCR, measuring α-glucosidase activity, the characterization of maltase gene copy number in deletion mutants using qPCR and flow cytometry. In context with the published data and results obtained in this study about the copy number of the maltase gene on C. utilis genome, we attempted to hypothesise and made conclusion about the ploidy of C. utilis. The results of this work, besides the biotechnological aspect, contribute to the elementary knowledge of C. utilis. The exact information about the ploidy or more specifically about the copy number of appropriate gene is essential for expression cassette dosage determination integrated into the chromosome of the host. In this study, we come to the conclusion that the maltase gene is present in C. utilis genome in four alleles, and in combination with flow cytometry, published information and the published genome sequences, the observations support the theory about tetraploidy of C. utilis.
Characterization of two types of yeast ribosomal DNA genes
Journal of bacteriology, 1978
The intragenic organization of ribosomal DNA from a diploid strain of Saccharomyces cerevisiae was analyzed by using recombinant DNA molecules constructed in vitro. Restriction analysis of the yeast ribosomal DNA with the EcoRI restriction enzyme indicated that eight restriction fragments were present in the ribosomal DNA of this strain: X' (1.87 X 10(6) daltons), A (1.77 X 10(6) daltons), B (1.48 X 10(6) daltons), C (1.22 X 10(6) daltons), D (0.39 X 10(6) daltons), E (0.36 X 10(6) daltons), F (0.22 X 10(6) daltons), and G (0.17 X 10(6) daltons). These fragments were distributed between two different types of ribosomal DNA genes, which had the restriction maps: (formula: see text) in which the underlined region shows the repeating unit. The diploid yeast strain contained approximately equal amounts of each of these two types of genes. The analysis of the recombinant DNA molecules also indicated that the yeast ribosomal genes are homogeneous and extensively clustered.