紫外诱变筛选低高级醇和双乙酰含量的啤酒酵母

根据酵母相关代谢途径，将降低高级醇和双乙酰含量为目的进行育种。采用紫外诱变方法对啤酒酵母进行处理，用乳酸平板、麦芽汁-碳酸钙平板、TTC上层平板进行显色分离，最终得到1株高级醇产量降低约30％、双乙酰含量亦较低的新菌株。     

激光诱变选育低双乙酰啤酒酵母

双乙酰是啤酒发酵过程中产生的一种重要的发酵副产物,也是衡量啤酒是否成熟的尺度。利用激光诱变选育双乙酰值低的啤酒酵母,得到一株优良菌株。用该菌株酿造的啤酒,其双乙酰值为0.1253mg/L。      

低双乙酰啤酒酵母激光诱变条件的研究

以激光为诱变剂照射啤酒酵母，得到1株双乙酰值低的啤酒酵母。用该菌株酿造的啤酒，发酵液中双乙酰含量为0．0836mg／L，比用原菌株酿造的啤酒下降了53％。激光诱变的条件为：波长623．8nm(红色)、功率6mW的He—Ne激光，用滤纸片法照射啤酒酵母，照射时间为5min。     

低双乙酰啤酒酵母的选育

将APV啤酒酵母菌株在不同的双乙酰含量梯度培养基内培养，挑出抗双乙酰的变异菌株，经菌种分离、筛选、发酵及双乙酰驯养等步骤选育出一株7^#菌，11℃低温发酵，双乙酰峰值0．36mg／L，成品啤酒双乙酰含量0．06mg／L，真正发酵度66．1％。     

紫外诱变及苯黄隆抗性处理选育低双乙酰啤酒酵母

将APV菌株经紫外线诱变后筛出苯黄隆抗性菌株,经实验室菌种分离、筛选、发酵及双乙酰驯养等步骤,筛选育出一株2#菌株,发酵双乙酰峰值为0.36mg/L,成品啤酒双乙酰含量为0.07 mg/L,真正发酵度为65.8%.     

两株酵母的双乙酰还原速度比较

本文比较了两种啤酒酵母菌株不同代数时的双乙酰还原速度,对嫩啤酒中的酵母细胞数与双乙酰的关系进行了研究。发现酵母菌株的不同或酵母代数的不同会导致双乙酰峰值不同,但还原速度相差不大。双乙酰的生成量与啤酒中的酵母细胞增殖密度成正比,双乙酰的还原速度与嫩啤酒中的酵母细胞数成正比。     

低双乙酰啤酒酵母工程菌的构建

利用PCR技术以啤酒酵母QY的染色体为模板扩增出含有乙酰羟酸合成酶（AHAS）基因ILV2的片段，将ILV2基因的内部EcoRI片段连接到整合载体YIp5上，并在该载体的Bam HI-SalI位点插入铜抗性基因CUP1－MT1，构建了YIpCE质粒，将其转化啤酒酵母QY，所得到的转化子AHAS酶的活力比受体菌QY降低75％左右， 在发酵测试中，转化了产生双乙酰的量比原始菌株降低30％。     

啤酒酵母原生质体制备与再生条件的筛选

试验正交设计方法,对啤酒酵母原生质体制备与再生进行了研究。试验表明:以静置培养12h的细胞,0.6mol/L KCl作为渗透压稳定剂,经EDTA-Na2+预处理,在2%蜗牛酶作用下,32℃酶解90min为条件获得最佳制备率,制备率为99%以上;以静置培养14h的细胞,0.6mol/L蔗糖作为渗透压稳定剂,不经过预处理,在1.5%蜗牛酶作用下,30℃酶解120min,涂布于山梨醇为渗稳剂的再生培养基中为条件获得最佳再生率,再生率为65%。     

原生质体紫外诱变技术在核糖菌种选育中的应用

本文采用原生质体紫外诱变技术,经过硫酸二乙酯(DES)诱变后筛选出的NO3菌。采用溶菌酶浓度0.8mg/ml,酶解温度36℃,酶解时间120min,获得原生质体;采用10s作为紫外原生质体诱变的作用时间,选育出一株稳定的核糖生产菌TJ3,该菌具有莽草酸营养缺陷,耐高糖、以核糖为唯一碳源不生长等遗传标记。     

啤酒酵母原生质体融合株GR8的中试研究

啤酒酵母原生质体融合株GR8的中试表明:融合株GR8凝絮性较强(本斯值为3.0);以12oBx麦芽汁为培养基,用500L发酵罐在12℃下发酵15d,发酵度为69.6%;发酵液中的双乙酰、乙醛、总高级醇、乙酸乙酯和乙酸异戊酯的含量分别为0.0390mg/L,3.26mg/L,84.7mg/L,11.14mg/L和0.99mg/L。     

固定化后期添加啤酒酵母菌降低啤酒中双乙酰含量的探讨

为探索用后期添加固定化酵母菌降低啤酒中双乙酰的工艺,通过单因素和正交试验,发现后期接入固定化啤酒酵母菌降低双乙酰含量的最佳工艺为:发酵温度为12℃,接入时间为发酵第16d,发酵再次接入固定化酵母菌的接种量为1.0%,此时双乙酰的量为0.083mg/L,啤酒口感达到9.2分。固定化的酵母菌可以重复利用3次,啤酒的双乙酰值和口感维持稳定。该研究表明利用固定化细胞的后期添加可以降低啤酒中的双乙酰含量。     

紫外线与亚硝基胍复合诱变选育高产海藻糖菌株的研究

本实验以酿酒酵母为出发菌株,用紫外线与亚硝基胍复合诱变选育出一株遗传稳定性较好的高产海藻糖菌株,其海藻糖含量为19.8%(以干酵母计),比原菌株提高了39.43%,比只用紫外线诱变后的菌株提高了10.0%,比只用亚硝基胍诱变后的菌株提高了6.45%。     

高产Monacolin K红曲霉的诱变选育及其与酿酒酵母共酵培养

为提高红曲中莫纳克林K含量,以实验室保藏紫色红曲霉（Monascus purpureus）MY-11为原始菌株,利用紫外诱变筛选红曲霉,并进行固态发酵制备红曲,采用高效液相色谱（HPLC）法对红曲中的莫纳克林K含量进行测定,选出高产莫纳克林K突变株,并与食源性菌株酿酒酵母（Saccharomyces cerevisiae）进行共酵。结果表明,通过紫外诱变筛选出2株高产莫纳克林K突变株M7和M8,固态发酵25 d后莫纳克林K含量分别为12.42 mg/g、12.49 mg/g,分别比原始菌株MY-11提高了26%、27%。将诱变菌株M7、M8分别继续与不同添加量的酿酒酵母共酵培养,结果发现加入酵母菌液添加量3%时莫纳克林K含量最高,分别为9.35 mg/g、8.94 mg/g,比对照组提高了2.63%、16.41%。紫外诱变筛选结合共酵培养方法可提高红曲中莫纳克林K含量。     

Isolation and characterization of barosensitive mutants of Saccharomyces cerevisiae obtained by UV mutagenesis

Using UV mutagenesis, 2 high-pressure (HP) sensitive (barosensitive) mutants of Saccharomyces cerevisiae were obtained. The HP inactivation of the mutants, as well as their parent strains, followed 1st-order kinetics in the range of 175 to 250 MPa within 600 s. Both mutants showed larger 1st-order inactivation rate constant values or significant loss of viabilities, compared with their parent strains in the pressure range tested. The inactivation rate constant value of one of the mutants was comparable with that of a previously reported highly barosensitive strain, which was generated by deletion of hsp104 in a trehalose deficient strain. The activation volume values of HP inactivation reactions in the 2 mutants were apparently equivalent with those of their parent strains. This suggested that the mutation did not bring drastic volume changes of the key molecules for HP inactivation. Their auxotrophic properties, growth, and ethanol fermentation were identical in mutant and parent strains. The mutants could therefore be useful for fermentations where control by HP processing is desired.     

原生质体复合诱变选育耐高温高产乙醇酵母菌株

以酿酒酵母Ygx-5为出发菌株,对其原生质体进行紫外线(UV)与亚硝基胍(NTG)复合诱变,经初筛和复筛,选育出1株耐高温、高产乙醇菌株U-N2。在37℃培养条件下,菌株U-N2产乙醇浓度最高可达16.32%vol,比原菌株提高20.44%,比只用紫外线诱变后的菌株提高8.04%,比只用亚硝基胍诱变后的菌株提高6.27%。经过20次传代培养,乙醇产量稳定。     

Red Beet Pigment Composition. Effects of Fermentation by Different Strains of Saccharomyces cerevisiae

Pasteurized red beet juice was inoculated with seven strains of Saccharomyces cerevisiae after adjusting pH to 5.8. Fermentation was at 26°C. The decay of red beet pigments was spectro-photometrically determined and expressed as betanine. Average percentages of betanine, isobetanine, betanidine, isobetanidine and the saccharide components were monitored by HPLC. After 40 hr fermentation, the strains Oenoferm, IO90 and Tokaj 76D showed pigment retentions and sucrose concentrations (g.L^-1) of 62% and traces, 58% and 3.4, and 56% and 1.4 respectively, and were considered the most suitable.     

Characterization of the rad14-2 Mutant of Saccharomyces cerevisiae: Implications for the Recognition of UV Photoproducts by the Rad14 Protein

The RAD14 gene of Saccharomyces cerevisiae is required for the incision step of the nucleotide excision repair process. The Rad14 protein can bind zinc, possesses a potential zinc finger DNA binding domain and has been shown to bind specifically to damaged DNA. Differences in UV sensitivity exist between a rad14 deletion strain and a putative rad14 point mutant, the point mutant being more resistant to UV than the deletion strain. Here, we confirm that the rad14 deletion strain repairs neither UV-induced cyclobutane pyrimidine dimers (CPDs) nor endonuclease III-sensitive damage sites, whereas the point mutant cannot repair the former but can repair the latter. From this it can be inferred that the point mutant produces an altered protein product allowing recognition of endonuclease III sensitive sites but not CPDs. To investigate this, the rad14 mutant allele was sequenced. It contained two GC-AT transition mutations when compared to the wild-type RAD14 gene sequence. When the rad14 point mutant sequence is translated, alterations within the putative zinc finger binding domain are observed, with one of the cysteine residues of the zinc binding motif being replaced by tyrosine. This suggests that alterations within the zinc finger binding domain of the Rad14 protein cause changes to the damage recognition properties of the protein. The use of the Rad14 protein from the point mutant should assist in experiments investigating the in vitro binding properties of the Rad14 protein to different types of DNA damage. © 1997 by John Wiley & Sons, Ltd.     

XV. Yeast sequencing reports. A gene from Saccharomyces cerevisiae which codes for a protein with significant homology to the bacterial 3-phosphoserine aminotransferase

During the sequencing of the gene GSP2 from Saccharomyces cerevisiae, we have encountered an adjacent open reading frame having strong homology to the 3-phosphoserine aminotransferase (E.C.2.6.1.52) from other organisms. In this report, we present the sequence for this yeast SERC, and evidence that its deletion from the yeast genome leads to serine dependency. The sequence has been deposited in the GenBank data library under Accession Number L20917.     

Subunit and cofactor binding of Saccharomyces cerevisiae sulfite reductase - towards developing wine yeast with lowered ability to produce hydrogen sulfide

Wine yeast, in common with other industrial Saccharomyces cerevisiae yeast, can produce hydrogen sulfide during alcoholic fermentation. The aim of this work was to evaluate a genetic strategy for reducing hydrogen sulfide production by lowering the activity of NADPH-dependent sulfite reductase, a key enzyme in the biosynthesis of the sulfur-containing amino acids. The role of amino acids predicted to be necessary for cofactor binding was investigated by comparisons with other proteins in the ferredoxin family. Substitutions in the glycine-rich loop domain (glycines 891 and 893) and at serine 953 of the alpha subunit, Met10p, appear to strongly affect NADPH binding. However, mutations in two amino acids that are completely conserved within the NADPH binding domain of the ferredoxin reductase family, lysine 959 and cysteine 987, have minimal effect on enzyme activity. Serine 820 is indicated to be essential for efficient FAD binding. The two-hybrid system was used to confirm that Met 1 0p interacts with Met5p, the predicted beta subunit of sulfite reductase. Binding of the subunits could only be demonstrated in the absence of methionine, indicating that an additional factor, possibly siroheme, is required to mediate this interaction. Strategies to decrease, but not eliminate, sulfite reductase activity in the cell using identified mutants were tested, and the findings are discussed with the a view to producing improved wine strains.