{"title":"Development of a real-time PCR protocol for the detection of chicken DNA in meat products.","authors":"Gulyaim Abitayeva, Arman Abeev","doi":"10.1080/10826068.2024.2317289","DOIUrl":"10.1080/10826068.2024.2317289","url":null,"abstract":"<p><p>Food falsification is a pressing issue in today's food industry, with fraudulent substitution of costly ingredients with cheaper alternatives occurring globally. Consequently, developing straightforward and efficient diagnostic systems to detect such fraud is a top priority in scientific research. The aim of the work was to develop a test system and protocol for polymerase chain reaction (PCR) to detect in food products of animal origin the substitution of expensive meat raw materials for by-products of poultry processing. For this, real-time polymerase chain reaction (RT-PCR) was used, which allows determining the qualitative and quantitative substitution in raw and technologically prepared products. Other methods for detecting falsification - enzyme immunoassay (ELISA/ELISA) or express methods in the form of a lateral flow immunoassay are less informative. The extraction of nucleic acids for real-time polymerase chain reaction depends on the source matrix, with higher concentrations obtained from germ cells and parenchymal organs. Extraction from muscle and plant tissues is more challenging, but thorough grinding of these samples improves nucleic acid concentration by 1.5 times using DNA extraction kits. The selection of primers and fluorescent probes through GenBank and PCR Primer Design/DNASTAR software enables efficient amplification and identification of target chicken DNA fragments in various matrices.</p>","PeriodicalId":20401,"journal":{"name":"Preparative Biochemistry & Biotechnology","volume":" ","pages":"1068-1078"},"PeriodicalIF":2.0,"publicationDate":"2024-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140102299","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Engineered <i>Saccharomyces cerevisiae</i> harbors xylose isomerase and xylose transporter improves co-fermentation of xylose and glucose for ethanol production.","authors":"Mengtian Huang, Xinxin Cui, Peining Zhang, Zhuocheng Jin, Huanan Li, Jiashu Liu, Zhengbing Jiang","doi":"10.1080/10826068.2024.2315479","DOIUrl":"10.1080/10826068.2024.2315479","url":null,"abstract":"<p><p><i>Saccharomyces cerevisiae</i> cannot assimilate xylose, second to glucose derived from lignocellulosic biomass. Here, the engineered <i>S. cerevisiae</i> strains INV<i>Sc</i>-XI and INV<i>Sc</i>-XI/XT were constructed using <i>xylA</i> and <i>Xltr1p</i> to co-utilize xylose and glucose, achieving economic viability and sustainable production of fuels. The xylose utilization rate of INV<i>Sc</i>-XI/XT was 2.3-fold higher than that of INV<i>Sc</i>-XI, indicating that overexpressing <i>Xltr1p</i> could further enhance xylose utilization. In mixed sugar media, a small amount of glucose enhanced the consumption of xylose by INV<i>Sc</i>-XI/XT. Transcriptome analysis showed that glucose increased the upregulation of acetate of coenzyme A synthetase (<i>ACS</i>), alcohol dehydrogenase (<i>ADH</i>), and transketolase <i>(TKL)</i> gene expression in INV<i>Sc</i>-XI/XT, further promoting xylose utilization and ethanol yield. The highest ethanol titer of 2.91 g/L with a yield of 0.29 g/g at 96 h by INV<i>Sc</i>-XI/XT was 56.9% and 63.0% of the theoretical ethanol yield from glucose and xylose, respectively. These results showed overexpression of <i>xylA</i> and <i>Xltr1p</i> is a promising strategy for improving xylose and glucose conversion to ethanol. Although the ability of strain INV<i>Sc</i>-XI/XT to produce ethanol was not very satisfactory, glucose was discovered to influence xylose utilization in strain INV<i>Sc</i>-XI/XT. Altering the glucose concentration is a promising strategy to improve the xylose and glucose co-utilization.</p>","PeriodicalId":20401,"journal":{"name":"Preparative Biochemistry & Biotechnology","volume":" ","pages":"1058-1067"},"PeriodicalIF":2.0,"publicationDate":"2024-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139723729","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Production of reverse transcriptase from Moloney murine Leukemia virus in <i>Escherichia coli</i> expression system.","authors":"Yudhi Nugraha, Fina Amreta Laksmi, Isa Nuryana, Helbert, Firyal Nida Khasna","doi":"10.1080/10826068.2024.2317311","DOIUrl":"10.1080/10826068.2024.2317311","url":null,"abstract":"<p><p>Reverse transcriptase (RT) is one of the most important enzymes used in molecular biology applications, enabling the conversion of RNA into complementary DNA (cDNA) that is used in reverse transcription-polymerase chain reaction (RT-PCR). The high demand of RT enzymes in biotechnological applications making the production optimization of RT is crucial for meeting the growing demand in industrial settings. Conventionally, the expression of recombinant RT is T7-induced promoter using IPTG in <i>Escherichia coli</i> expression systems, which is not cost-efficient. Here, we successfully made an alternative procedure for RT expression from Moloney murine leukemia virus (M-MLV) using autoinduction method in chemically defined medium. The optimization of carbon source composition (glucose, lactose, and glycerol) was analyzed using Response Surface Methodology (RSM). M-MLV RT was purified for further investigation on its activity. A total of 32.8 mg/L purified M-MLV RT was successfully obtained when glucose, glycerol, and lactose were present at concentration of 0.06%, 0.9%, and 0.5% respectively, making a 3.9-fold improvement in protein yield. In addition, the protein was produced in its active form by displaying 7462.50 U/mg of specific activity. This study provides the first step of small-scale procedures of M-MLV RT production that make it a cost-effective and industrially applicable strategy.</p>","PeriodicalId":20401,"journal":{"name":"Preparative Biochemistry & Biotechnology","volume":" ","pages":"1079-1087"},"PeriodicalIF":2.0,"publicationDate":"2024-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139973159","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Cynthia Lizbeth López-García, Guadalupe Guerra-Sánchez, Fortunata Santoyo-Tepole, Dario Rafael Olicón-Hernández
{"title":"Chitinase induction in <i>Trichoderma harzianum</i>: a solid-state fermentation approach using shrimp waste and wheat bran/commercial chitin for chitooligosaccharides synthesis.","authors":"Cynthia Lizbeth López-García, Guadalupe Guerra-Sánchez, Fortunata Santoyo-Tepole, Dario Rafael Olicón-Hernández","doi":"10.1080/10826068.2024.2313631","DOIUrl":"10.1080/10826068.2024.2313631","url":null,"abstract":"<p><p>This study innovatively employed solid-state fermentation (SSF) to evaluate chitinase induction in <i>Trichoderma harzianum</i>. Solid-state fermentation minimizes water usage, a crucial global resource, and was applied using shrimp waste chitin and a mixture of commercial chitin with wheat bran as substrates. Shrimp waste and wheat bran were pretreated and characterized for SSF, and the fungus's utilization of the substrates was assessed using spectrophotometric and microscopic methods. The resulting enzymes' ability to produce chitooligosaccharides (COS) mixtures was studied. Wheat bran/commercial chitin demonstrated superior performance, with a 1.8-fold increase in chitinase activity (76.3 U/mg protein) compared to shrimp waste chitin (41.8 U/mg protein). Additionally, the COS mixture obtained from wheat bran/commercial chitin showed a higher concentration of reducing sugars, reaching 87.85 mM, compared to shrimp waste chitin (14.87 mM). The COS profile from wheat bran/commercial chitin included monomers to heptamers, while the profile from shrimp waste chitin was predominantly composed of monomers. These results highlight the advantages of SSF for chitinase induction and COS production in <i>T. harzianum</i>, offering potential applications as dietary fiber, antioxidants, and antimicrobial agents. The findings contribute to by-product valorization, waste reduction, and the sustainable generation of valuable products through SSF-based enzyme production.</p>","PeriodicalId":20401,"journal":{"name":"Preparative Biochemistry & Biotechnology","volume":" ","pages":"1040-1050"},"PeriodicalIF":2.0,"publicationDate":"2024-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139723726","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Muinat Moronke Adeyanju, Adedeji Nelson Ademakinwa
{"title":"Tyrosinase from the pulps of local cultivars of <i>Musa spp:</i> Purification, characterization, immobilization, and application in the batch production of l-3,4-dihydroxyphenylalanine.","authors":"Muinat Moronke Adeyanju, Adedeji Nelson Ademakinwa","doi":"10.1080/10826068.2024.2324084","DOIUrl":"10.1080/10826068.2024.2324084","url":null,"abstract":"<p><p>Tyrosinase, an enzyme involved in browning reactions in plants/crops exposed to mechanical injury, was isolated from the pulp of some different locally available bananas (<i>M. cavendish</i>, <i>M. acuminata</i>, and <i>M. paradisiaca</i>). Tyrosinase from the pulps was extracted, purified, immobilized, and characterized. Thereafter, the potentials of the immobilized tyrosinase in the possible production of l-3,4-dihydroxyphenylalanine (L-DOPA) in an improvised batch reactor was exploited using tyrosine and ascorbate as the substrates. L-DOPA production was monitored via thin-layer chromatography and spectrophotometry (Arnow's method). L-DOPA is a drug that is used in the treatment of Parkinson's disease. Hence, this study exploited a non-chemical route for its synthesis using the tyrosinase obtained from the banana pulps. The purified tyrosinase had an optimum pH and temperature of 6.5 and 7.0, respectively. The molecular weight of the purified tyrosinase was 45 kDa. Quercetin and resorcinol both competitively inhibited the purified tyrosinase from the three cultivars. Immobilized <i>M. cavendish</i> tyrosinase produced the highest concentration (0.60 mM) of L-DOPA after 8 h in an improvised batch reactor. The tyrosinase in the banana pulps serves as a cheap and readily available green route for the possible production of L-DOPA.</p>","PeriodicalId":20401,"journal":{"name":"Preparative Biochemistry & Biotechnology","volume":" ","pages":"1098-1105"},"PeriodicalIF":2.0,"publicationDate":"2024-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140040158","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Modification of media using food-grade components for the fermentation of <i>Bifidobacterium</i> and <i>Lactobacillus</i> strains in large-scale bioreactors.","authors":"Chayanee Boontun, Savitri Vatanyoopaisarn, Sungwarn Hankla, Eisuke Kuraya, Yasutomo Tamaki","doi":"10.1080/10826068.2020.1861009","DOIUrl":"10.1080/10826068.2020.1861009","url":null,"abstract":"<p><p>Probiotic bacteria continue to receive increasing attention in the food and feed industries. However, the production of <i>Bifidobacterium</i> and <i>Lactobacillus</i> at an industrial scale is challenging because of specific nutrient requirements and conditions, which are complicated and costly. We developed low-cost culture media by modifying the carbon and nitrogen sources for <i>Bifidobacterium animalis</i> subsp. <i>lactis</i> KMP-H9-01 and <i>Lactobacillus reuteri</i> KMP-P4-S03 from available food grade components. Sucrose (15 g/l) was selected as a suitable carbon source for both strains because it was the most economic and facilitated bacterial growth that was equal to that of glucose. The <i>Bifidobacterium</i> strain required beef extract as a nitrogen source to multiply. The fermentation of both strains using the modified media formula in 5-L and 50-L bioreactors showed that the highest cell counts of <i>L. reuteri</i> and <i>B. animalis</i> subsp. <i>lactis</i> were 9 and 9.8 log CFU/ml after 12-15 h, respectively. The concentration (g/l) ratio between lactate and acetate obtained from <i>B. animalis</i> subsp. <i>lactis</i> was 7:7.4 at 12 h and 11.4:10.6 at 40 h; the ratio was similar at both time points (6.9: 1.1) for <i>L. reuteri</i>. Thus, this economically modified food-grade medium for the large-scale fermentation of two probiotic bacteria was efficient.</p>","PeriodicalId":20401,"journal":{"name":"Preparative Biochemistry & Biotechnology","volume":" ","pages":"1017-1027"},"PeriodicalIF":2.0,"publicationDate":"2024-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"38785445","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Recombinant expression and purification of phenylalanyl-tRNA synthetase from wheat: a long-lasting poly(U)-dependent poly(Phe) synthesis system.","authors":"Haruyuki Furukawa, Yuto Nagashio, Kensuke Tsutsumi, Naofumi Matsubara, Ryohei Kato, Chie Tomikawa, Kazuyuki Takai","doi":"10.1080/10826068.2024.2324077","DOIUrl":"10.1080/10826068.2024.2324077","url":null,"abstract":"<p><p>Synthetic genes for the two subunits of phenylalanyl-tRNA synthetase (PheRS) from wheat were expressed in <i>Escherichia coli</i>. When each gene was induced individually, the α subunit with a cleavable 6 × His tag at the amino terminus was largely soluble, while the β subunit was almost completely insoluble. When the two subunits were co-expressed, a soluble fraction containing the two subunits were obtained. This was purified by a standard method in which the tag was cleaved off with a specific protease after affinity purification. As the sample contained slightly more PheRSα than PheRSβ, we further resolved the sample by gel filtration to obtain the fraction that showed the size of the conventional <i>α</i><sub>2</sub><i>β</i><sub>2</sub> tetrameric complex and contains an almost equal amount of the two subunits. The final yield was 0.6 mg per 1 liter of the culture medium, and the specific activity was 28 nmol min<sup>-1 </sup>mg<sup>-1</sup>, which was higher than that of a fraction purified from wheat germ. This recombinant PheRS was used, along with purified samples of the elongation factors and the ribosomes from wheat germ, for a poly(U)-dependent poly(Phe) synthesis reaction. The reaction was dependent on the added components and lasted for more than several hours.</p>","PeriodicalId":20401,"journal":{"name":"Preparative Biochemistry & Biotechnology","volume":" ","pages":"1088-1097"},"PeriodicalIF":2.0,"publicationDate":"2024-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140028774","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Production, purification, properties and current perspectives for modification and application of microbial lipases.","authors":"Junxin Zhao, Maomao Ma, Zheling Zeng, Dongman Wan, Xianghui Yan, Jiaheng Xia, Ping Yu, Deming Gong","doi":"10.1080/10826068.2024.2323196","DOIUrl":"10.1080/10826068.2024.2323196","url":null,"abstract":"<p><p>With the industrialization and development of modern science, the application of enzymes as green and environmentally friendly biocatalysts in industry has been increased widely. Among them, lipase (EC. 3.1.1.3) is a very prominent biocatalyst, which has the ability to catalyze the hydrolysis and synthesis of ester compounds. Many lipases have been isolated from various sources, such as animals, plants and microorganisms, among which microbial lipase is the enzyme with the most diverse enzymatic properties and great industrial application potential. It therefore has promising applications in many industries, such as food and beverages, waste treatment, biofuels, leather, textiles, detergent formulations, ester synthesis, pharmaceuticals and medicine. Although many microbial lipases have been isolated and characterized, only some of them have been commercially exploited. In order to cope with the growing industrial demands and overcome these shortcomings to replace traditional chemical catalysts, the preparation of new lipases with thermal/acid-base stability, regioselectivity, organic solvent tolerance, high activity and yield, and reusability through excavation and modification has become a hot research topic.</p>","PeriodicalId":20401,"journal":{"name":"Preparative Biochemistry & Biotechnology","volume":" ","pages":"1001-1016"},"PeriodicalIF":2.0,"publicationDate":"2024-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140040130","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Recent advancements in soluble expression of recombinant antibody fragments in microbial host systems.","authors":"Preeti Saroha, Rucha S Patil, Anurag S Rathore","doi":"10.1080/10826068.2024.2394446","DOIUrl":"https://doi.org/10.1080/10826068.2024.2394446","url":null,"abstract":"<p><p>Recombinant fabs dominate the pharmaceutical pipelines today with microbial host systems continuing to be a major contributor toward their production. <i>Escherichia coli</i> is a versatile host for recombinant protein expression due to its simplicity, affordability, and ability to be cultivated at high cell density. It is particularly suitable for non-glycosylated proteins and small proteins. Despite the aforementioned benefits, the use of <i>E. coli</i> as the host for the synthesis of recombinant antibody fragments often suffers from low yield and reduced activity. In most cases, proteins are expressed as inclusion bodies and need to undergo refolding to achieve their active forms and this refolding step is generally low-yielding. In this article, we review the various approaches that researchers have taken to enhance the production of recombinant antibody fragments in <i>E. coli</i>. Molecular biology-oriented approaches such as cloning, chaperone-mediated folding, and host cell screening as well as process optimization involving examination of process parameters, media, and feeding have been addressed.</p>","PeriodicalId":20401,"journal":{"name":"Preparative Biochemistry & Biotechnology","volume":" ","pages":"1-10"},"PeriodicalIF":2.0,"publicationDate":"2024-08-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142086245","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}