Isolation of lactoperoxidase, lactoferrin, α-lactalbumin, β-lactoglobulin B and β-lactoglobulin A from bovine rennet whey using ion exchange chromatography

A mild and rapid method is described for isolating various milk proteins from bovine rennet whey. β-Lactoglobulin from bovine rennet whey was easily adsorbed on and desorbed from a weak anion exchanger, diethylaminoethyl-Toyopearl. However, α-lactalbumin could not be adsorbed onto the resin. α-Lactalbumin and β-lactoglobulin from rennet whey could also be adsorbed and separated using a strong anion exchanger, quaternary aminoethyl-Toyopearl. The rennet whey was passed through a strong cation exchanger, sulphopropyl-Toyopearl, to separate lactoperoxidase and lactoferrin. α-Lactalbumin and β-lactoglobulin were adsorbed onto quaternary aminoethyl-Toyopearl. α-Lactalbumin was eluted using a linear (0–0.15 M) concentration gradient of NaCl in 0.05 M Tris–HCl buffer (pH 8.5). Subsequently, β-lactoglobulin B and β-lactoglobulin A were eluted from the column with 0.05 M Tris–HCl (pH 6.8), using a linear (0.1–0.25 M) concentration gradient of NaCl. The yields were 1260 mg α-lactalbumin, 1290 mg β-lactoglobulin B and 2280 mg β-lactoglobulin A from 1 l rennet whey.     

The use of limited proteolysis to probe interdomain and active site regions of beta-galactosidase (Escherichia coli)

Limited proteolysis by pancreatic elastase (EC 3.4.21.36) and chymotrypsin (EC 3.4.21.1) was used to study the domain structure and active site of beta-galactosidase (EC 3.2.1.23) (Escherichia coli). Treatment with elastase resulted in a rapid cleavage between residues Ala-732 and Ala-733. No inactivation accompanied this cleavage suggesting that this bond is in a hinge region of the protein. Some slow cleavages beyond the initial one were observed to occur and were accompanied by inactivation. Treatment of beta-galactosidase with chymotrypsin resulted in cleavages first between Trp-585 and Ser-586 and then between Phe-601 and Cys-602. The first of these cleavages resulted in total inactivation of beta-galactosidase. The presence of monovalent ions or isopropyl-beta-D-thiogalactopyranoside protected against the cleavages but when Mg2+ or Mn2+ was present in the reaction mixture, the bond between Trp-585 and Ser-586 was more susceptible to the action of chymotrypsin. These data demonstrate that the conformation of beta-galactosidase around Trp-585 and Ser-586 is dramatically affected by the binding of ions and isopropyl-beta-D-thiogalactopyranoside. The mutant M15 beta-galactosidase, which is missing residues 11 through 41 and is an inactive dimer rather than an active tetramer, was found to be much more labile to proteases than native beta-galactosidase, but the same initial cleavages were found to occur. In addition, trypsin cleaved the M15 protein between Arg-431 and Trp-432 while native beta-galactosidase was stable to trypsin.     

糙叶败酱挥发油镇静作用的研究

本文观察了败酱科植物糙叶败酱(Patrinia scabra Bunge)根和根茎中制得的挥发油的镇静作用,井与黄花败酱挥发油做了比较。结果表明,此油灌胃给予数组小鼠,剂量0.45ml/kg,显示如下的作用:[1]能显著延长由于腹腔注射戊巴比妥钠(50mg/kg)引起的小鼠睡眠时间,但其作用强度弱于黄花败酱挥发油。(2)一次灌胃给予小鼠大剂量10.46g/kg的糙叶败酱挥发油,连续观察10天,动物外观正常,无一死亡,体重增加与对照组相似。     

鲢、鳙在天然条件下的摄食强度(Ⅱ)武汉东湖鲢、鳙周年摄食强度的研究

武汉东湖的鲢(Hypophthalmichthy molitrix)鳙(Aristichthys nobilis)在天然条件下摄食强度具季节性变化。摄食强度高峰处于夏季,低谷处于冬季。在实验条件下,按周年采样期间水温变化范围,测定鱼的肠管排空率。食物通过鱼肠管时间(Y_p—h)与水温(X_t—℃)的关系为: 鲢Y_p=270.63 X_t~(0.6408) 鳙Y_p=280.46 X_t~(0.6642) 根据修正后Bajkov公式(D=C (24.A)/n),估算鱼的日粮。鱼日粮(Y_D)与水温(X_t)关系为: 鲢Y_D=0.2683e~(0.1503X_t) 鳙Y_D=0.0075X_t~(2.2715) 计算鱼在天然条件下周年月粮及年粮。鲢、鳙对天然饵料年消耗量分别为18.924公斤及17.39公斤,饵料系数分别为18.02及13.38。     

“引黄济青”输水沿线水生生物分布与蓄水库富营养化关系的研究

“引黄济青”是国家“七五”重点工程,由黄河下游博兴县境内打渔张闸引水,在渠首沉沙后,再经惠民、东营、潍坊和青岛的10个县区约253km的明渠输送,到达唠山县棘洪滩蓄水库蓄存,实现向青岛市日供水3.0×10~5m~3。明渠输水过程中,来白沿线的营养物质会污染水体,加之输水在库中滞留时间长,会不会发生富营养化,这是社会普遍关注的问题。     

质粒YRP7的基本特性鉴定

质粒YRP7用氯霉素法扩增,碱变性裂解法提取,酸酚法及核糖核酸酶纯化后,得到了高产量(5.6mg/L培养液),高纯度(A260:A280=2.0)的质粒制品,经转化实验及酶切分析确定YRP7具有下列特征:大小为5.41±0.10kb,可赋予宿主细胞AmP~r、Tet~r的表型,对大肠杆菌C600的转化频率为10~(-6)、转化效率为1.5×10~6转化子/mgDNA。限制性内切酶BamH Ⅰ、ECoRⅠ、Hind Ⅲ及PstⅠ在其分子上的切点数分别为1、2、2、2,并确定了各酶切片段的分子大小,对BanHⅠ的单切点,经插入失活法证实其位于Tet~r的基因上。由上述特征可确定,质粒YRP7是一个比较理想的克隆载体。     

可育的抗除草剂溴苯腈转基因小麦

报道了采用微粒轰击（Ｍｉｃｒｏｐｒｏｊｅｃｔｉｌｅ ｂｏｍ ｂａｒｄｍ ｅｎｔ） 幼胚将除草剂抗性基因导入小麦（Ｔｒｉｔｉｃｕｍａｅｓｔｉｖｕｍ Ｌ．）的转化研究。实验共使用了１３ 个小麦品种, 从开花后１４～１８ ｄ 的籽粒中剥取幼胚, 植物表达质粒含有ＣａＭＶ ３５Ｓ启动子控制的除草剂溴苯腈抗性基因ｂｘｎ 以及筛选标记基因ＮＴＰⅡ。采用高压放电基因枪,用质粒ＤＮＡ 包被的钨粒轰击预培养３ ｄ 的幼胚。在含有卡那霉素类似物ｇｅｎｅｔｉｃｉｎ Ｇ４１８ｓｕｌｐｈａｔｅ 的ＭＳ培养基上, 经过多步骤筛选和分化, 从８００ 多个幼胚中获得了１６ 株转化苗。除草剂抗性鉴定和Ｓｏｕｔｈｅｒｎ 杂交分析证明, 其中４ 株为转基因植物,具有溴苯腈抗性, 并且自交可育。转化工作从分离幼胚到转化苗鉴定完毕, 最短时间为６ 个月, 因此, 该方法是一项快速有效的基因导入技术