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The sequence diversity and expression among genes of the folic acid biosynthesis pathway in industrial Saccharomyces strains

100%
Goncerzewicz A., Misiewicz A.
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issue 4
841-850
EN
Folic acid is an important vitamin in human nutrition and its deficiency in pregnant women's diets results in neural tube defects and other neurological damage to the fetus. Additionally, DNA synthesis, cell division and intestinal absorption are inhibited in case of adults. Since this discovery, governments and health organizations worldwide have made recommendations concerning folic acid supplementation of food for women planning to become pregnant. In many countries this has led to the introduction of fortifications, where synthetic folic acid is added to flour. It is known that Saccharomyces strains (brewing and bakers' yeast) are one of the main producers of folic acid and they can be used as a natural source of this vitamin. Proper selection of the most efficient strains may enhance the folate content in bread, fermented vegetables, dairy products and beer by 100% and may be used in the food industry. The objective of this study was to select the optimal producing yeast strain by determining the differences in nucleotide sequences in the FOL2, FOL3 and DFR1 genes of folic acid biosynthesis pathway. The Multitemperature Single Strand Conformation Polymorphism (MSSCP) method and further nucleotide sequencing for selected strains were applied to indicate SNPs in selected gene fragments. The RT qPCR technique was also applied to examine relative expression of the FOL3 gene. Furthermore, this is the first time ever that industrial yeast strains were analysed regarding genes of the folic acid biosynthesis pathway. It was observed that a correlation exists between the folic acid amount produced by industrial yeast strains and changes in the nucleotide sequence of adequate genes. The most significant changes occur in the DFR1 gene, mostly in the first part, which causes major protein structure modifications in KKP 232, KKP 222 and KKP 277 strains. Our study shows that the large amount of SNP contributes to impairment of the selected enzymes and S. cerevisiae and S. pastorianus produce reduced amounts of the investigated metabolite. The results obtained here yield a list of genetically stable yeast strains which can be implemented as a starter culture in the food industry.
2
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Expression of GPD1 and SIP18 genes during rehydration in active dry industrial Saccharomyces cerevisiae cider-making yeast strains (ADY)

81%
Goncerzewicz A., Kamińska-Wojteczek K., Młynarczyk I., Misiewicz A.
Acta Biochimica Polonica
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2017
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vol. 64
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issue 2
287-294
EN
In this study we determined the influence of different sugar concentration in media, time of rehydration and type of strain on relative expression level of GPD1 and SIP18 genes of active dry cider-making yeast strains, followed by the assessment of the impact of rehydration on the fermentation process. High expression of SIP18 at the beginning of rehydration was shown to be due to high transcription of the gene during the drying process. High sugar concentrations of media initiated transcription of the GPD1 gene and triggered the cellular glycerol biosynthesis pathway in examined strains. Rehydration time and type of strain showed to have no statistically significant impact on the course of the fermentation; RT qPCR results depended mainly on the time of rehydration and sugar concentration of the medium. This is the first attempt to confront rehydration time and molecular mechanisms acting upon rehydration with the course of the fermentation process.
3
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Intra-strains diversity of expression of polymorphic PKS4 gene in comparison in zearalenone production by Fusarium graminearum during in vitro cultivation

81%
Misiewicz A., Goncerzewicz A., Jędrzejczak R., Zdziennicki F.
Acta Biochimica Polonica
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2016
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vol. 63
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issue 1
97-102
EN
Filamentous fungi belonging to the Fusarium genus are responsible for large economic losses due to their high pathogenicity and toxigenicity. Fusarium sp. may produce variety of mycotoxins, one of them is zearalenone (ZEA). The presence of the PKS4 gene shows the possibility of zearalenone biosynthesis by Fusarium sp. In this study, in four Fusarium graminearum and one Fusarium poae strains the presence of PKS4 genes and ZEA concentrations were determined. The presence of the PKS4 gene was confirmed by classical polymerase chain reaction (PCR) in three of four strains of F. graminearum. One strain with no PKS4 gene detected was found while still producing ZEA. In the present study, a real-time PCR assay has been successfully performed for the relative expression of Fusarium strains based on new designed primers targeting the PKS4 gene involved in ZEA biosynthesis. Result shows that P56/4 strain of F. graminearum has the highest mRNA level, in the range of 12, what correlates to the high production of this mycotoxin. In this study, a real-time PCR assay has been successfully developed for the prediction of the production of ZEA by F. graminearum strains by PCR real-time techniques based on primers targeting the gene, PKS4, involved in ZEA biosynthesis. The special significance was pointed to occurring genes polymorphism.
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