Effects of Plant Extracts and Beauveria bassiana on the Activity of Defense-Related Enzymes in Solanum lycopersicum L.during Interaction with Fusarium oxysporum f.sp.lycopersici

The objective to this work was to evaluate the enzymatic activity in the culture of Solanum lycopersicum L.infected with Fusarium oxysporum after the combined application of Beauveria bassiana and plant extracts.Solanum lycopersicum plantlets were transplanted 15 days after the emergency.Five days after transplanting,Beauveria bassiana spores were applied at a concentration of 1×10^(7)spores mL^(−1)onto soil(along with A.indica(N)and P.auritum(H)leaf extracts)where S.lycopersicum plants were planted.Eight days after transplanting,spores of F.oxysporum strain were applied at a concentration of 1×10^(6)spores mL^(−1)to soil where S.lycopersicum plants were growing.The development of S.lycopersicum plants was monitored for 114 days,whereby a randomized complete block treatment design was used,the roots of the plants were crushed with liquid nitrogen,after which UV/VIS spectrophotometry was used to determine protein concentration,as well as the activity ofβ-1,3-glucanases,peroxidases(POX,EC1.11.1.7),catalase and chitinases.The treatments combining B.bassiana and A.indica and P.auritum extracts,had a significant difference in enzymatic activity levels,thus contributing to better defense mechanisms and a greater protection to S.lycopersicum plants in the presence of F.oxysporum.The application of B.bassiana and plant extracts to the ground mitigated damage caused by F.oxysporum on S.lycopersicum plants.

采用酵母双杂交法筛选PSRPK相关蛋白基因

本课题组在前期研究中,从多头绒泡菌中分离到一个SR蛋白激酶基因psrpk,并将其编码蛋白命名为PSRPK(SR protein kinase ofPhysarum Polycephalum).为分离PSRPK相关蛋白基因以了解PSRPK的功能,构建了转化率为2×106转化子/3μg pGADT7-Rec、密度为5.35×108cells/mL、滴度为2.34×109cfu/mL的多头绒泡菌酵母双杂交AD库.以PSRPK为饵蛋白筛选该文库得到接合率为41.18%的杂交酵母,在SD/-Leu/-Trp/-Ade/-H is培养板上筛选获得1476个杂交克隆,其中,X-gal滤纸显色呈强蓝色的克隆有342个.对大于500 bp的67个克隆测序获得35个cDNA片段,其中编码Plasm in C、branched-chain am inoacid am inotransferase(BCAT)类似蛋白、MSF1类似蛋白、m ixed-linked glucanase precursor类似蛋白、FCY1p类似蛋白和40S ribosomal protein S2类似蛋白等7个cDNA片段在阳性杂交酵母克隆中出现多次,其余仅出现一次.在35个cDNA片段的编码序列中有15个具有同源蛋白,31个编码序列的丝氨酸含量大于或等于6%,符合激酶底物的组成特征.

Characterization of Exo 1, 4-<i>β</i>glucanase produced from <i>Tricoderma Viridi</i>MBL through solid-state bio-processing of orange peel waste

Agro-industrial residues are primarily composed of complex polysaccharides that strengthen the microbial growth for the production of industrially important enzymes like cellulases. In the present study we aimed to characterize the Exo 1, 4-β glucanase that was indigenously produced from Trichoderma viride MBL. T. viride MBL was cultured in the Solid-State medium of orange peel (50% w/w moisture) under optimized fermentation conditions and maximum activity of 412 ± 12 U/mL was recorded after 4th day of incubation at pH 5.5 and 30℃. Exo 1, 4-β glucanase was 4.17-fold purified with specific activity of 642 U/mg in comparison to the crude extract. To confirm its purity and molecular weight, sodium dodecyl sulphate poly acrylamide gel electrophoresis (SDS- PAGE) was performed. The enzyme was shown to have a molecular weight of 60 kDa with an optimum pH and temperature of 5 and 50℃, respectively. Lineweaver-Burk reciprocal plot revealed that the kinetic constants Km and Vmax of purified Exo 1, 4-β glucanase were 76 μM and 240 U/mL.

Effects of dietary barley inclusion and glucanase supplementation on the production performance,egg quality and digestive functions in laying ducks

This study evaluated the effects of barley inclusion and glucanase supplementation on the productive performance and digestive function in laying ducks.The experiment used a randomized design with a 5×2 factorial arrangement of 5 graded levels of barley(0%,15%,30%,45%and 60%)with or without 1.5 g/kgβ-1,3-1,4-glucanase(15,000 U/kg).During the experimental period of 120 d,the weight and total number of eggs within each pen were recorded daily,and egg quality was determined every 4 wk.At the end of the experiment,3 randomly selected ducks within each replicate were sacrificed,then duodenal digesta and jejunal mucosa was collected.Dietary inclusion of barley had no effects on egg production,daily egg mass or FCR,but supplementation with glucanase improved egg production and FCR(P<0.01).Barley did not affect feed intake of laying ducks,but glucanase tended to increase feed intake(P=0.09).Neither barley norβ-glucanase had effects on the egg quality variables,except for yolk color score,which was decreased with increasing barley supplementation.Glucanase,but not barley,increased the activity of chymotrypsin and amylase in duodenal digesta.Barley inclusion affected the activity of alkaline phosphatase and maltase in jejunal mucosa(P<0.05),butβ-glucanase had no effects on the activity of these brush border enzymes.Barley inclusion increased the glucan content in duodenal digesta,but supplementation of glucanase to barley-based diet reduced digesta glucan content and reduced total volatile fatty acids and increased the proportion of acetic acid in cecal contents.The results indicate that,without glucanase,the optimal dietary barley level in the diets of laying ducks is about 13%for maximal production performance;glucanase supplementation of the barley diets improved production perfor-mance,probably through enhancing digestive function.