产植酸酶枯草芽孢杆菌诱变选育方法的研究

通过对产植酸酶枯草芽孢杆菌进行不同方法的诱变处理,确定最佳诱变条件.对产植酸酶的枯草芽孢杆菌进行筛选,然后采用紫外(UV)、亚硝基胍(NTG)、UV-NTG和NTG-UV4种方法对其进行诱变.UV诱变(100 s)选育的Z3菌株植酸酶活性最高(71.96 U/mL),比筛选出发菌株提高38.46％.结果表明:UV (100 s)选育方法为最佳的诱变条件.     

利用原生质体诱变筛选植酸酶高产菌株

对产植酸酶的纯化黑曲霉用混合酶制取原生质体进行了研究。研究发现,经诱变后的原生质体在再生过程中菌落形态发生了明显变化,从中选育出了一株植酸酶的高产菌株,并对其遗传稳定性进行了鉴定。     

具有万古霉素高产能的东方拟无枝酸菌诱变株的选育研究

为提高万古霉素(Vancomycin)发酵生产水平,选育出其发酵生产的优良菌株,以东方拟无枝酸菌(Amycolatopsis oriertalsis)菌株V1806作为原始菌株,利用EMS(甲基磺酸乙酯)、UV(紫外诱变)和ARTP(常压室温等离子体诱变)等单一诱变、复合诱变的方法,选育出1株遗传稳定性好、且对万古霉素耐受性强的菌株Vua-15,其中试发酵效价达到7863 mg/L,比原始菌株提高了74.3%。研究结果表明,复合诱变对东方拟无枝酸菌菌种选育的效果优于单一诱变,更易选育出符合预期的优良菌株。研究不仅对万古霉素工业化发酵生产具有重要意义,而且为其他产品发酵菌种的选育提供了一定借鉴。     

蛋鸡日粮中添加粗制植酸酶的经济和社会效益分析

植酸酶虽然能解除植物性饲料中植酸的抗营养作用,降低无机磷源的添加量,减轻环境污染,但目前市场上的植酸酶价格偏高,成为阻碍植酸酶被普遍使用的主要原因。为此,本课题组利用自然界中的黑曲霉菌,经60Co辐射形成诱变菌株,从中选出产酶能力较高的A菌株,以价格低廉的麸皮、棉子粕、菜子粕为基质进行固体发酵,并在生产中以粗酶的形式添加,以期降低植酸酶的使用成本。本实验室以前的毒性试验和饲养试验表明,此粗制植酸酶无毒副作用,在蛋鸡日粮中可取代一部分的无机磷。本试验着重分析在蛋鸡日粮中使用此粗制植酸酶的经济价值和社会价值。一、材…     

螺旋霉素高产菌株的选育

以螺旋霉素链霉菌(Streptomyces spiramyceticus)SPM-14078菌株作为出发菌株,利用常压室温等离子体(ARTP)诱变、紫外线(UV)诱变和亚硝基胍(NTG)诱变的方法对菌株进行诱变处理,比较3种方法的诱变效果,通过诱变筛选出1株遗传稳定性好且具有Co~(2+)和豆油耐受性、噬菌体抗性的菌株SPMAR-1603,其发酵效价比出发菌株提高了294.9%,此结果对螺旋霉素高效生产、品质提升具有重要意义。     

不同诱变方法对TL-15028菌株产泰乐菌素相关性能的影响

为了选育出泰乐菌素高产菌株,以弗氏链霉菌(Streptomyces fradiae)TL-15028菌株作为出发菌株,利用化学诱变、物理诱变以及复合诱变的方法对菌株进行诱变处理,比较不同诱变方法的诱变效果,通过诱变筛选出1株遗传稳定性好、且具有泰乐菌素和豆油抗性的菌株TLEU-1503,其发酵效价比出发菌株提高了36.5%,达到14124μg/m L,对泰乐菌素高效生产、品质提升具有重要意义。     

不同诱变方法对TL-15028菌株产泰乐菌素相关性能的影响

为了选育出泰乐菌素高产菌株,以弗氏链霉菌(Streptomyces fradiae)TL-15028菌株作为出发菌株,利用化学诱变、物理诱变以及复合诱变的方法对菌株进行诱变处理,比较不同诱变方法的诱变效果,通过诱变筛选出1株遗传稳定性好、且具有泰乐菌素和豆油抗性的菌株TLEU-1503,其发酵效价比出发菌株提高了36.5%,达到14124μg/m L,对泰乐菌素高效生产、品质提升具有重要意义。     

中性蛋白酶高产菌株的选育

试验采用米曲霉(Aspergillus oryzae)菌株作为出发菌株,希望通过诱变处理,选育高产菌株,以达到工业化生产的需要。通过筛选和产酶试验,选育出的菌株P10,产中性蛋白酶酶活可达2000U/g,,比出发菌株提高了67%,产酶性能稳定,能够适应规模化生产的需要,实现了试验的预期目标。1材料和方法1.1材料1.1.1菌株现有的米曲霉菌株以及经诱变后纯培养得到的53个菌株,编号为A1-A28及P1-P25。1.1.2主要仪器与设备分析天平、恒温水浴锅、分光光度计、温度计、秒表、高压灭菌锅、超净工作台等。1.1.3主要试剂和溶液福林试剂、0.4mol/LNa2CO3、0.4mol/LC…     

常用诱变育种技术在我国真菌育种上的应用

<正>自然选育是获得优良菌株最基本的方法,从自发突变的菌株中选择优良菌株,这种方法被一直沿用下来,由于自发突变率较低,要从自然界筛选到理想的菌株,需要很长的时间,花费很大的人力财力,在科技飞速发展的今天已经满足不了需要。诱变育种则补充了自然选育的不足,能够在较短的时间内,通过改变微生物的遗传特性,获得能够满足需要的理想菌株。真菌诱变育种,是以人工诱变手段诱发真菌基因     

紫外诱变选育黄霉素高产菌

采用紫外线诱变方法对黄霉素产生菌进行诱变处理,利用菌株抗自身代谢物的特征,在添加高浓度黄霉素的分离培养基上选育获得比出发菌株高的突变株sg2-U-5,效价比原始菌株提高了5.4倍。而且突变株的菌落形态与亲本菌株差异较大。     

黑曲霉液态发酵产植酸酶的条件优化研究

文章以黑曲霉为出发菌株,通过初筛、复筛,得到一株产植酸酶的菌株,对发酵时间、温度及pH进行单因素试验,并利用正交设计进行工艺优化。结果表明,当培养基pH为5.5、温度为30℃、发酵时间为5d时,黑曲霉发酵产植酸酶活力最大,为14.35U/mL。     

Effects of a rice diet and phytase addition on growth performance,tissue weights,phosphorus and nitrogen retention,and on liver threonine dehydrogenase,malic enzyme and fatty acid synthase activities in broiler chicks

This experiment was conducted to clarify the nutritional functions of rice and phytase addition for broiler chicks. Thirty‐six 7‐day‐old male chicks (ROSS 308 strain) were assigned to one of the four treatment groups: corn‐ or rice‐based diet groups and each diet with added phytase (2000 phytase units/kg diet) groups (corn + P or rice + P groups). The non‐phytate phosphorus (npP) content in the diets with added phytase was approximately half of the requirement. Body weight gain and feed intake in the rice group was significantly higher than those in the corn group. Breast and thigh muscle weights and nitrogen retention in the rice group were significantly higher than that in the corn group. Although the efficiency of phosphorus retention (%) in the corn + P group was significantly higher than that in the corn group, no significant difference was observed between the rice and rice + P groups. Liver threonine dehydrogenase activity in the corn group was significantly higher than in the other three groups. These results indicate that rice is superior to corn as a starter diet in broiler chicks, and that phytase action in the rice‐based diet was less than that in the corn‐based diet.     

Efficacy of a genetically modified yeast phytase on phosphorus bioavailability in a corn-soybean meal based diet for growing pigs

We investigated the efficacy of genetically modified yeast (GMY) phytase on phosphorus (P) bioavailability in growing pigs fed a corn–soybean meal based diet. Crossbred barrows were fed a P-adequate, a low-P or a P-deficient diet containing 0.37, 0.27 and 0.14% non-phytate-P, respectively. The P-deficient diet was supplemented with wild-type yeast (WTY), Aspergillus (ASP) or GMY phytase at 750 PU per kilogram of food. Dietary ASP and GMY phytases increased plasma inorganic P (P_i) concentration and the apparent absorption of P, and decreased the concentration of a bone resorption marker, plasma type-I collagen C-terminal telopeptide (ICTP). Wild-type yeast phytase also increased the apparent absorption of P, but the changes in plasma P_i and ICTP concentrations were not significant. Although the dietary P_i-equivalencies based on plasma P_i and ICTP concentrations did not differ between WTY and ASP phytases, the equivalency of ASP phytase based on apparent absorption of P was higher than that of WTY phytase. The equivalency of GMY phytase calculated from each parameter was higher than that of WTY phytase, and did not differ from that of ASP phytase. These results suggest that efficacy of GMY phytase on P bioavailability was higher than WTY phytase, and as effective as ASP phytase in growing pigs.     

Effects of Broiler Strain,Dietary Nonphytate Phosphorus,and Phytase Supplementation on Chick Performance and Tibia Ash

Three experiments were conducted to study the effects of broiler strain and phytase supplementation on chick nonphytate P (NPP) requirements for growth, feed intake, and tibia ash. The first experiment compared the NPP requirements for 8- to 22-d-old chicks from 2 broiler strains, Ross 308 and 708, that have been selected for differences in early weight gain and performance. The second experiment utilized similar 8- to 22-d-old Ross 308 and 708 chicks but also compared the effects of dietary fungal phytase supplementation (600 U/kg) on broiler NPP requirements. The third experiment utilized a younger starting age, 5 to 23 d old, for Ross 308 and 708 chicks with and without phytase supplementation. Minor differences in chick growth did not affect chick NPP requirements in Experiments 1 and 3, but a substantial and unexplained reduction of growth of the Ross 708 chicks in Experiment 2 resulted in a lower NPP requirement for chick growth and feed intake but not for tibia ash. As expected, supplementation of diets with fungal phytase did result in decreased NPP requirements for growth, feed intake, and tibia ash in both strains used in Experiment 3.     

Efficacy of yeast phytase in improving phosphorus bioavailability in a corn-soybean meal-based diet for growing pigs

Crossbred barrows (n = 66; 6 wk old) were used in a 6-wk experiment to evaluate the efficacy of phytase from yeast or Aspergillus niger on performance, tibial characteristics, and serum inorganic P concentration. We also investigated the stability of these phytases in acidic solutions with pepsin, which simulated gastric conditions. Pigs were fed a P-adequate diet containing .34% nonphytate-P or a low-P diet containing .20% nonphytate-P. The low-P diet was supplemented with 0, 1,000, 2,000, or 4,000 phytase units (PU; the activity at optimal pH, i.e., pH 4.2 for yeast phytase and pH 5.5 for phytase from Aspergillus niger)/kg of yeast phytase, or 1,000 PU/kg phytase from Aspergillus niger. The graded level of yeast phytase linearly increased ADG (P = .047), tibial weight (P = .091), tibial density (P < .001), and P concentration in tibial cortex (P = .018). Aspergillus niger phytase also increased ADG (P = .022), serum inorganic P concentration (P < .001), tibial density (P = .007), and tibial P concentration (P = .025). The pigs given 1,000 PU/kg Aspergillus niger phytase showed greater ADG (P = .091), tibial density (P= .001), and tibial P concentration (P = .062) than those given 1,000 PU/kg yeast phytase. No measurements differed (P > .31) between the pigs given 1,000 PU/kg Aspergillus niger phytase and those given 4,000 PU/kg yeast phytase. These results suggested that yeast phytase improves bioavailability of P in the diet for growing pigs but the efficacy of yeast phytase is less than that of Aspergillus niger phytase. During incubation in acidic solutions with pepsin, yeast phytase (P < .001) lost more of its activity than Aspergillus niger phytase. This lesser stability of yeast phytase may be responsible for the poorer efficacy of yeast phytase than that of Aspergillus niger. In summary, supplementation of swine diets with yeast phytase is beneficial, but its efficacy is less than that of Aspergillus niger phytase.     

Brewer's yeast efficiently degrades phytate phosphorus in a corn-soybean meal diet during soaking treatment

Microbes such as yeast and Aspergillus are known to produce phytase, and Aspergillus phytase has been used as a feed additive for improving phytate-phosphorus bioavailability in monogastric animals. We measured phytase activity in some by-products from fermented food and beverage productions by yeast and Aspergillus . The phytase activity was as high as 3577 and 2225 PU/kg DM in raw and dried brewer's yeasts, respectively. On the other hand, the phytase activity was approximately 400 PU/kg DM in white-wine yeast and red-wine yeast. The phytase activity was further low in natto (fermented soybean) residue, soy sauce cake, rice brewer's grain and the activity was not detected in dried corn-barley distiller's grain with soluble and sweet-potato distiller's residue. The stability of phytase against pepsin was much lower in the brewer's yeast than in an Aspergillus phytase preparation. On the other hand, the addition of raw brewer's yeast effectively degraded phytate phosphorus in a corn-soybean meal diet during soaking. These results suggest that phytase in the examined by-products is not suitable for the phytase source of conventional diets, but that the soaking treatment with a raw brewer's yeast is an alternative method for improving phytate-phosphorus bioavailability in corn-soybean meal diets for pigs.     

Feed formulation using phytase in laying hen diets

Lohmann LSL-Lite and Classic Brown laying pullets were reared and caged at the Agassiz Research Centre following recommendations of the management guides. At 30 wk of age, these hens were fed 1 of 5 diets per strain following the Lohmann nutrient recommendations. In addition to a control treatment, diets were formulated to contain 300 or 600 U/kg of phytase, with or without enzyme inclusion, where the phytase was assumed to cause the release of P, Ca, energy, and protein. Diets were changed at 45 wk to follow a phase-feeding program, but treatments remained the same. These dietary changes did not result in major changes in measures associated with P deficiency, likely because the management guides suggest P levels that largely exceed the requirements of the birds. Rather than adding high levels of inorganic P to layer feeds, the safety margin currently included in recommended dietary specifications could be provided by the addition of phytase. This would reduce the negative environmental effects of intensive poultry production that are associated with P excretion.     

不同清洁型日粮降低规模猪场中氮、磷污染物排泄的研究

本试验旨在研究在肥育猪日粮中添加植酸酶、复合酶及微生态制剂3种清洁型日粮对规模猪场氮、磷排泄的影响。试验选取健康无病的60 kg左右的苏钟猪15头,随机分为5个处理,分别为对照组(正常磷水平)、负对照组(不加CaHPO₄)、负对照+植酸酶组、对照+植酸酶+复合酶+和美酵素组、对照+微生态制剂组。结果表明,3种清洁型日粮均可降低粪、尿中氮的排泄量,其中植酸酶组粪中氮的排泄量较对照组降低了0.55%(P＞0.05),植酸酶+复合酶+和美酵素组降低了10%(P＜0.05),微生态制剂组降低了6.11%(P＞0.05),其中植酸酶+复合酶+和美酵素组减排效果更佳,尿氮含量各组之间差异不显著(P＞0.05);粪中磷的排泄量植酸酶组较对照组降低了19.19%(P＞0.05),植酸酶+复合酶+和美酵素组降低了36.63%(P＜0.05),微生态制剂组降低了16.28%(P＞0.05),其中植酸酶+复合酶+和美酵素组减排效果最佳,尿磷的排泄量植酸酶组降低了12%(P＞0.05),植酸酶+复合酶+和美酵素组降低了16%(P＜0.05),微生态制剂组降低了12%(P＞0.05),以植酸酶+复合酶+和美酵素组减排效果最好。由此表明,在肥育猪日粮中添加清洁饲料可有效降低规模猪场中氮、磷排泄造成的环境污染,且以植酸酶、复合酶及和美酵素联合应用效果最佳。     

日粮、食糜中植酸酶活性的测定

本试验以2～10周龄的海兰褐蛋鸡为研究对象，用玉米 豆粕为基础日粮，设1个对照组和6个处理组。通过对比7组日粮，研究日粮、食糜中的植酸酶活性。结果表明：日粮中植酸酶活性随着麸皮添加量的增加，逐渐增强。但肌胃食糜中未检测出酶活，而小肠食糜检测出酶的活性。并且随着鸡只日龄的增加，小肠食糜的酶活逐渐增强。     

Comparative enzymatic hydrolysis of phytate in various animal feedstuff with two different phytases

Bacillus amyloliquefaciens DS11 phytase (DS11 phytase) and Aspergillus ficuum phytase (AF phytase) activities were investigated by measuring the release of phosphate from phytate in animal feedstuff such as wheat bran, corn meal, soybean meal and rice flour at pH 5 and 7. In all the tested feedstuff, the enzymatic activity of DS11 phytase was more active at pH 7, but that of AF phytase was more active at pH 5. From these results, the phytate in the gastrointestinal tract could be degraded in the small intestine or stomach by DS11 or AF phytase, respectively. In conclusion, the results presented in this paper indicated that different combination ratios of DS11 and AF phytase, depending on the kind of feedstuff, might effectively induce more enzymatic activity both in the stomach and small intestine in terms of the pH of the gastrointestinal tract.