利用MPCR方法快速检测植物转基因背景

利用复合ＰＣＲ（Ｍｕｌｙｉｐｌｅｘ ＰＣＲ－ＭＰＣＲ）技术对植物的转基因背景进行检测。经过对ＤＮＡ方法的选择,对各种ＰＣＲ程序的比较以及对引物的修饰,建立了一种快速检测植物转基因情况的技术;利用该技术对５个大豆样品,６个豆粕样品进实验检测,同时利用普通ＰＣＲ方法对上述样品进行检测,两者的结果完全符合。     

“大豆分离蛋白”工艺用复合酶制剂的研究

大豆分离蛋白的碱浸提酸沉淀工艺,目前普遍存在蛋白质回收率低,分离蛋白成品得率低,纯度低的问题,利用复合酶制剂的作用可改变这一状况。针对工艺需要,从十余种工业酶制剂中筛选出两种蛋白酶活低、多糖降解酶活高的酶制剂;Ａｎ－７６半纤维酶和果胶酶ｕｌｔｒａｚｙｒｍ＾ＴＭ。对两种酶制剂的酶活组分、主体酶的部分酶学性质,以及酶的用量与蛋白质浸出率的关系等方面进行了大量基础实验。经优化实验确定了复合酶制剂的复配比     

Covalent immobilization of alkaline proteinase on amino-functionalized magnetic nanoparticles and application in soy protein hydrolysis

Magnetic nanoparticles (MNPs) were synthesized and surface modified with (3-Aminopropyl)triethoxysilane (APTES). The alkaline proteinase (AP) was covalently immobilized on the APTES-modified MNPs through glutaraldehyde linkage. The resulting AP-loaded MNPs have an average size of 84 nm in aqueous solution, and a magnetization of 40 emu/g, endowing the immobilized enzyme with excellent magnetic responsively and dispersity. The maximum amount of AP and catalytic activity immobilized 1.0 mg MNPs was 120 μg and 25.3 units, respectively. Immobilized AP showed maximum activity at pH 10.0 and 50°C. Compared with free enzyme, the immobilized AP exhibited better storage stability. Moreover, immobilized AP can be reused 10 times and still maintained about 50% of its initial activity. The degree of hydrolysis of soy protein hydrolysates for immobilized AP could reach 19.0%, which was closer to the value of free enzyme. The molecular weight (M.W.) analysis showed that the soy protein was hydrolyzed successfully into small peptides of two main fractions with an average M.W. of 742 and 2126 Da. This study indicated that the immobilized AP could be used to hydrolyze continuously soy protein for potential industry application. © 2018 American Institute of Chemical Engineers Biotechnol. Prog., 35: e2756, 2019.     

EFFICIENT PROTEOLYSIS AND APPLICATION OF AN ALKALINE PROTEASE FROM HALOPHILIC Bacillus sp. EMB9

A salt-stable alkaline protease from moderately halophilic Bacillus sp. EMB9, isolated from the western coast of India, is described. This protease was capable of efficiently removing silver from used/waste X-Ray films, as well as hydrolyzing defatted soy flour with 31% degree of hydrolysis (DH). Production of the protease was optimized by using response surface methodology. Ca²⁺ and NaCl were the most critical factors in enhancing the yield. Under optimized culture conditions, a maximum of 369 U protease/mL was obtained, which is quite comparable to the yields of commercial proteases. The elevated production level coupled with ability to efficiently hydrolyze protein-laden soy flour and complete recovery of silver from used X-Ray films makes it a prospective industrial enzyme.     

Diets containing pomegranate polyphenol and soy isoflavone attenuate contact hypersensitivity in mice

Contact hypersensitivity (CHS) is frequently used as an animal model for human allergic contact dermatitis (ACD). Diets of pomegranate polyphenols (PPs) or soy isoflavones (SIs) each alleviated CHS symptoms; however, the effect of diets containing a mixture of PPs and SIs on CHS is unclear. We investigated the CHS-inhibitory effects of diets supplemented with a mixture of PPs and SIs at human physiologically relevant doses. Consuming the mixture of PPs and SIs attenuated ear swelling and reduced infiltration of Gr-1-positive cells. Ear swelling decreased in the PP and SI-treated mice compared to the SI-treated mice. The auricle tissues of the PP and SI-fed mice exhibited decreased production of CXCL2 and MCP-5 compared to the SI- and PP-treated mice, respectively. These results suggest that dietary supplementation with a mixture of PPs and SIs may have ACD-preventive effects and may prove more beneficial than supplementation with PPs or SIs alone.     

Characterization of Tn3926, a new mercury-resistance transposon from Yersinia enterocolitica

A new transposon coding for mercury resistance (HgR), Tn3926, has been found in a strain of Yersinia enterocolitica, YE138A14. The element has a size of 7.8 kb and transposes to conjugative plasmids belonging to different incompatibility groups. A restriction map has been established. DNA-DNA hybridization indicates that Tn3926 displays homology with both Tn501 and Tn21; the greatest homology is shown with the regions of these transposons that encode HgR. Weaker homology is observed between Tn3926 sequences and those regions of Tn501 and Tn21 that encode transposition functions. Complementation experiments indicate that the Tn3926 transposase mediates transposition of Tn21, albeit somewhat inefficiently, but not of Tn501, while the resolvase mediates resolution of transposition cointegrates formed via Tn21, Tn501, or Tn1721.     

Characterization of soy protein hydrolysates and influence of its iron content on monoclonal antibody production by a murine hybridoma cell line

A challenging aspect with the use of protein hydrolysates in commercial manufacturing processes of recombinant therapeutic proteins is their impacts on the protein production due to a lack of understanding of batch-to-batch variability. Soy hydrolysates variability and its impact on fed-batch production of a recombinant monoclonal antibody (mAb) expressed in Sp2/0 cells were studied using 37 batches from the same vendor. The batch-to-batch variability of soy hydrolysates impacted cell growth, titer and product quality. Physicochemical characterization of batches confirmed that soy hydrolysates are mainly a source of amino acids and peptides containing lower amounts of other components such as carbohydrates and chemical elements in cell culture media. Soy hydrolysates composition of different batches was consistent except for trace elements. Statistical analyses identified iron as a potential marker of a poor process performance. To verify this correlation, two forms of iron, ferric ammonium citrate and ferrous sulfate, were added to a batch of soy hydrolysates associated to a low level of iron during cell culture. Both forms of iron reduced significantly cell growth, mAb titer and increased level of the acidic charge variants of the mAb. Consequently, trace element composition of soy hydrolysates or of all incoming raw materials might lead to significant impacts on process performance and product quality and therefore need to be tightly controlled.