瘤胃真菌分离鉴定及其在青贮饲料中应用效果评价
发布时间：2019-05-17 21:09
【摘要】：农业生物质资源是我国极其丰富的、可再生的生物燃料,化工原料和饲料来源。然而,由于农作物秸秆细胞壁多糖成分的复杂性、合成酶系的多样性和超分子结构的异质性导致其难以被畜禽有效利用,造成极大的浪费。厌氧真菌具有很强的降解植物组织的能力,是草食动物消化道内一类重要的降解植物细胞壁的功能菌。本文以西农萨能奶山羊瘤胃真菌为研究对象,利用亨盖特厌氧滚管技术,分离获得高植物细胞壁降解酶活性厌氧真菌,并研究其作为添加剂在全株玉米青贮中的应用效果,全面评价瘤胃厌氧真菌降解植物细胞壁功效及应用潜力,为今后瘤胃真菌资源的开发及在饲料工业的应用提供理论依据和技术支持。高植物细胞壁降解酶活性瘤胃真菌分离鉴定及酶学特性研究本研究采用亨盖特厌氧滚管技术从西农萨能奶山羊瘤胃中分离获得12株具有细胞壁降解酶活性的厌氧真菌,通过形态学观察、核糖体内转录间隔区和28S rDNA D1/D2区基因序列分析确定其分类地位。测定12株真菌的植物细胞壁降解酶活性(木聚糖酶、羧甲基纤维素酶、微晶纤维素酶、乙酰酯酶和β-葡聚糖酶),并对活性最高菌株的木聚糖酶和乙酰酯酶进行酶学特性分析。结果表明,12株瘤胃厌氧真菌均为Piromyces属,分别命名为Piromyces sp.CN1~Piromyces sp.CN12。其中,Piromyces sp.CN6的木聚糖酶、羧甲基纤维素酶和乙酰酯酶活性分别为1655.3、93.4和152.8 mU,显著高于其他菌株(P0.05),Piromyces sp.CN3的微晶纤维素酶活性最高,但与Piromyces sp.CN6差异不显著(P0.05),各菌株间β-葡聚糖酶活性差异不显著(P0.05)。木聚糖酶与羧甲基纤维素酶、乙酰酯酶存在极显著正相关(P0.01),与微晶纤维素酶呈显著正相关(P0.05)。酶学特性表明,木聚糖酶最适反应温度为50℃,最适pH为5.0,该酶在40℃和pH 5.0~8.0下较稳定;K+、Co2+、Ca2+对其有激活作用,Zn2+、Cu2+、Mg2+、Fe2+、Mn2+抑制该酶活性。乙酰酯酶的最适反应温度为50℃,最适pH为9.0,该酶在40℃和pH 5.0~10.0下较稳定;Mg2+、K+、Ca2+对乙酰酯酶有激活作用,Zn2+、Fe2+、Co2+、Mn2+抑制该酶活性。本研究从西农萨能奶山羊瘤胃内容物分离获得12株Piromyces属厌氧真菌,Piromyces sp.CN6具有较高的植物细胞壁降解酶活性,其中木聚糖酶和乙酰酯酶在较宽的温度和pH范围内均具有较高的活性,同时具有良好的热稳定性和酸碱稳定性。Piromyces sp.CN6对全株玉米青贮发酵品质、营养成分及体外降解率的影响本研究旨在验证试验一所筛选的高植物细胞壁降解酶活性瘤胃真菌Piromyces sp.CN6作为青贮添加剂对全株玉米青贮发酵品质、营养成分及体外降解率的影响。试验采用单因素完全随机设计,每组设5个重复,以全株玉米为青贮原料,分别设为对照组、真菌组(105 TFU/g)和复合酶组(0.033 mg/g),聚乙烯袋真空包装,室温下存储10 d、30 d、60 d后开封取样。结果表明,真菌组和复合酶组青贮饲料达到1级优良,对照组达到2级较好,瘤胃真菌Piromyces sp.CN6作为青贮接种剂能够在青贮前期大量繁殖并附着在玉米秸秆茎叶表面。与对照组相比,真菌组和复合酶组均可显著降低发酵30 d青贮饲料pH、乙酸含量和NH3-N/TN(P0.05),显著降低发酵10 d、30 d、60 d青贮饲料NDF和ADF含量(P0.05),显著提高发酵30 d青贮饲料可溶性碳水化合物含量、粗蛋白含量、乳酸含量和乳酸/乙酸比值(P0.05)。与对照组相比,真菌接种剂可显著提高发酵30 d青贮饲料DM、NDF和ADF体外降解率(P0.05)。不同处理对青贮饲料干物质回收率无显著影响(P0.05)。综上,瘤胃真菌Piromyces sp.CN6能够改善青贮饲料的发酵品质和营养成分,提高粗纤维的降解率。本研究从西农萨能奶山羊瘤胃分离获得12株Piromyces属厌氧真菌,其中Piromyces sp.CN6具有较高的植物细胞壁降解酶活性,且各酶之间具有较高的相关性。酶学特性分析表明:Piromyces sp.CN6分泌的木聚糖酶和乙酰酯酶在较宽的温度和pH范围内均具有较高的活性,同时具有良好的热稳定性和酸碱稳定性。瘤胃厌氧真菌能够改善青贮饲料的发酵品质和营养成分,提高粗纤维的降解率。
[Abstract]:The agricultural biomass resource is a rich and renewable bio-fuel, chemical raw material and feed source in China. However, due to the complexity of the polysaccharide component of the cell wall of the crop straw, the diversity of the synthetase system and the heterogeneity of the supramolecular structure cause it to be difficult to be effectively utilized by the livestock and poultry, resulting in great waste. The anaerobic fungus has strong ability to degrade plant tissue, and is an important functional bacterium for degrading plant cell wall in the digestive tract of herbivorous animal. In this paper, the rumen fungi of the nongaseng milk goat in the west of this article are the research objects, and the high plant cell wall degrading enzyme active anaerobic fungus is obtained by using the Henligate anaerobic rolling tube technology, and the application effect of the anaerobic fungus as an additive in the whole plant corn silage is studied. The effects of the rumen anaerobic fungi on the plant cell wall and the application potential are comprehensively evaluated, and the theoretical basis and technical support are provided for the development of the rumen fungus resources and the application of the feed industry in the future. In that present study,12 strain of anaerobic fungus with cell wall-degrading enzyme activity were obtained from the rumen of the western agricultural sacha milk goat by using the high-plant cell wall-degrading enzyme-active rumen fungus separation and identification and the enzyme-study property. The classification status of the ribosomal internal transcribed spacer region and the 28S rDNA D1/ D2 region gene sequence analysis was determined. The activity of plant cell wall-degrading enzymes (xylanase, methylcellulose, microcrystalline cellulose, and B-glucanase) of 12 strains of fungi was determined, and the xylanase and glyoxesterase of the highest active strain were analyzed. The results showed that the 12 strains of the rumen anaerobic fungi were Piromytsp. CN1 to Piromytsp. CN12, respectively. Among them, the activity of xylanase of Piromytsp. CN6 was 1655.3, 93.4 and 152.8mU, which was higher than that of other strains (P0.05). There was no significant difference in the activity of the E-glucanase between the strains (P0.05). There was a significant positive correlation between the xylanase and the methyl cellulose (P 0.01), and it was positively correlated with the microcrystalline cellulose (P <0.05). The optimum reaction temperature of xylanase is 50 鈩，
本文编号：2479409
[Abstract]:The agricultural biomass resource is a rich and renewable bio-fuel, chemical raw material and feed source in China. However, due to the complexity of the polysaccharide component of the cell wall of the crop straw, the diversity of the synthetase system and the heterogeneity of the supramolecular structure cause it to be difficult to be effectively utilized by the livestock and poultry, resulting in great waste. The anaerobic fungus has strong ability to degrade plant tissue, and is an important functional bacterium for degrading plant cell wall in the digestive tract of herbivorous animal. In this paper, the rumen fungi of the nongaseng milk goat in the west of this article are the research objects, and the high plant cell wall degrading enzyme active anaerobic fungus is obtained by using the Henligate anaerobic rolling tube technology, and the application effect of the anaerobic fungus as an additive in the whole plant corn silage is studied. The effects of the rumen anaerobic fungi on the plant cell wall and the application potential are comprehensively evaluated, and the theoretical basis and technical support are provided for the development of the rumen fungus resources and the application of the feed industry in the future. In that present study,12 strain of anaerobic fungus with cell wall-degrading enzyme activity were obtained from the rumen of the western agricultural sacha milk goat by using the high-plant cell wall-degrading enzyme-active rumen fungus separation and identification and the enzyme-study property. The classification status of the ribosomal internal transcribed spacer region and the 28S rDNA D1/ D2 region gene sequence analysis was determined. The activity of plant cell wall-degrading enzymes (xylanase, methylcellulose, microcrystalline cellulose, and B-glucanase) of 12 strains of fungi was determined, and the xylanase and glyoxesterase of the highest active strain were analyzed. The results showed that the 12 strains of the rumen anaerobic fungi were Piromytsp. CN1 to Piromytsp. CN12, respectively. Among them, the activity of xylanase of Piromytsp. CN6 was 1655.3, 93.4 and 152.8mU, which was higher than that of other strains (P0.05). There was no significant difference in the activity of the E-glucanase between the strains (P0.05). There was a significant positive correlation between the xylanase and the methyl cellulose (P 0.01), and it was positively correlated with the microcrystalline cellulose (P <0.05). The optimum reaction temperature of xylanase is 50 鈩，
本文编号：2479409
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