一种提取花生种子中总RNA的简易方法

介绍了一种适合花生种子的RNA提取方法。它具有简便、快速等特点,且所提取的RNA样品产率高,质量好,符合分子生物学实验要求。     

花生、大豆、芝麻与绿茶复合饮料的研制

以花生、大豆、白芝麻、绿茶叶、姜为主要原料,添加适量的其它辅料,经烘烤、浸泡、磨浆、调配、均质、脱气、杀菌等工艺研制出口感优良、风味独特、香气浓郁、营养丰富、质量稳定的具有一定保健功效的饮料.试验采用对比试验、正交试验找出最佳工艺条件和配方.该饮料的最佳配方为:100mL饮料中混合浆液50mL,姜茶汁20mL,糖6g,柠檬酸0.15g.     

一种RNA提取试剂盒--TRIZOL的使用方法初探

以酿酒酵母为原料，利用TRIZOL试剂快速提取法，得到的总RNA不完整，只有5s rRNA一条条带，经过对该试剂盒的改进后提取的总RNA呈现28s rRNA，18s rRNA和5s rRNA三条清晰的条带，很少有降解，其A260／A280值可达1．946，A260／A230值为2．070，具有很高的纯度。     

纯芝麻酱中花生致敏蛋白Ara h2、Ara h3及芝麻蛋白2S albumim的分析鉴定比较

针对花生致敏蛋白Ara h2、Ara h3及芝麻蛋白2S albumim,使用实时荧光定量PCR法(Real-time PCR)及酶联免疫吸附法(ELISA)对纯芝麻酱中上述蛋白进行检测.两种方法的特异性、重复性方法学验证显示均良好.验证后使用实时荧光定量PCR法对纯芝麻酱DNA进行芝麻蛋白2S albumim基因、花...     

大豆卵磷脂DNA的提取方法及应用

以大豆卵磷脂为研究对象,比较了CTAB法、磁珠试剂盒法以及硅胶膜试剂盒法等3种DNA提取纯化方法。通过大豆内源基因Lectin实时荧光PCR检测,发现CTAB法提取的大豆卵磷脂DNA质量优于其它两种方法,并以CTAB法为基础进行条件优化,进一步提高DNA纯度。采用该方法对6个不同品牌的市售大豆卵磷脂产品进行转基因成分(CAMV35S启动子及NOS终止子)检测,结果证实均不含转基因成分。     

一种从荞麦子叶中提取总RNA的有效方法及其RT-PCR检测

从植物组织中提取高质量的RNA是进行植物分子生物学研究的必要前提和关键,本研究以长出第一对子叶的荞麦为材料,采用CTAB-LiCl法获得荞麦总RNA。消除了蛋白质、DNA、多糖、多酚等污染。该方法提取荞麦子叶总RNA,完整性好、纯度高,检测结果说明所得总RNA产品可作RT-PCR、RACE等进一步的实验之用,此法无需特殊试剂和贵重仪器,实验结果稳定,重复性好,适合富含多糖和脂质的植物组织总RNA的提取。     

DNA 提取方法对实时荧光定量 PCR 检测花生过敏原 Ara h 2 基因的比较研究

目前,食物过敏是一个世界性的公共卫生问题,其中花生过敏最为严重。基于DNA的分子生物学检测方法目前已广泛应用于花生过敏原检测,样品DNA的提取质量会显著影响检测的灵敏度及准确率。本研究比较了5种DNA提取方法,包括十六烷基三甲基溴化铵（cetyl trimethyl ammonium bromide,CTAB）法、柱式法及3种基于磁珠纯化技术的DNA快速提取法,对生花生、煮花生、炸花生、烤花生、花生酥和花生酱6种不同加工方式的花生基质样品进行DNA提取,考察了不同方法获得的DNA浓度、纯度指标,并采用实时荧光定量PCR（quantitative real-time PCR, qPCR）对花生过敏原Ara h 2基因进行了检测分析。结果表明,月桂酰肌氨酸钠（Sodium Lauroyl Sarcosine）磁珠法（简称SLS磁珠法）的适用性广、提取率高,对于6种花生基质提取的DNA均能高效检出花生过敏原Ara h 2基因;对于花生含量为0.05%～1.00%的小麦粉二元混合物,SLS磁珠法的DNA提取率总体优于CTAB法,并且能有效提取出与花生共用一条生产线的燕麦片中污染的花生DNA,证实SLS磁珠法提取的实际样品DNA能够满足花生过敏原检测目的。本研究为花生及其制品DNA提取方法提供了参考,特别是磁珠类方法,高效快速,提取质量能够保障后续基于DNA的花生过敏原分子生物学方法检测结果的准确性。     

食品中甲型肝炎病毒RNA提取方法的比较及应用

目的：对食品中甲型肝炎病毒的3 种RNA提取方法进行综合比较,以优选出最佳的核酸提取方法,为各级实验室开展食源性甲型肝炎病毒核酸检验提供技术参考。方法：分别采用ABI AM1836、QIAGEN 74104和 ROCHE11858882001三种商业化核酸提取试剂盒对人工污染甲型肝炎病毒的食品样品进行核酸提取,根据3 种核酸提取方法的提取效率、抑制剂去除效率、检测灵敏度、综合应用指标进行病毒RNA的优化,且采用优化后的提取方法进行实际样品的检测。结果：综合病毒各项评价指标显示,ROCHE 11858882001在食品中提取甲型肝炎病毒RNA效果最优,其次为QIAGEN 74104和ABI AM1836。101 份样品的检测结果显示,一份蓝靛果样品为甲型肝炎病毒核酸阳性,其余样品均为阴性。结论：ROCHE 11858882001是一种快捷有效的病毒RNA提取方法,适用于食品中甲型肝炎病毒的日常检测工作。     

大豆胚芽油三种提取方法的比较研究

采用索氏提取法、超声波强化提取法和超临界CO2萃取法分别提取大豆胚芽油。在对提取工艺优化的基础上，将3种方法的萃取效率及所得胚芽油的理化性质、磷脂、不皂化物含量和脂肪酸组成进行分析比较，探讨萃取方法与油品的关系，全面评价萃取方法的优劣。研究结果表明：索氏提取法的出油率最高，但提取时间长；超声波强化法的出油率次之，提取时间最短，但溶剂用量大；超临界CO2萃取法出油率较低，但时间较短，流程简单无溶剂残留。超临界CO2萃取法所得胚芽油的磷脂含量低，不皂化物及不饱和脂肪酸含量高。综合考虑，超临界CO2萃取法是提取大豆胚芽油的最佳方法。     

4种豆类植物种子油脂的提取与分析

以常见的4种豆子为原料,粉碎后采用超声波辅助提取刀豆、花豆、大豆、红豆中的油脂,测定4种豆类植物种子中油脂的得率、理化性质、成分与含量。结果表明:除大豆外,其他3种豆类得油率较低,但是4种豆油在理化性质上都具有较好品质,符合食用植物油标准;在油脂成分上,4种豆油不饱和脂肪酸总量均较高,刀豆油中不饱和脂肪酸总量达到了91.43%,但4种豆油中大豆油含量最高的成分是亚油酸,而其他3种豆油含量最高的成分却是油酸,刀豆油中棕榈油酸含量相当高,达到16.2%,且亚麻酸含量也较高。     

Diminution of quinolizidine alkaloids,oligosaccharides and phenolic compounds from two species of Lupinus and soybean seeds by the effect of Rhizopus oligosporus

Lupinus, like Glycine max (soybean), is an ancient leguminous plant. It has been used as a food by people living around the Mediterranean Sea and in the Andean Highlands. This legume contains quinolizidine alkaloids (Qas), oligosaccharides (OGS) and phenolic compounds (PC). The objective of this study was to evaluate the effect of the growth of Rhizopus oligosporus on the levels of Qas, OGS and PC from L. mutabilis, L. campestris and G. max through tempeh elaboration. The results showed that the soaking and cooking processes of legume seeds diminished the Qas content of L. mutabilis and L. campestris by 50%, and after 48 h of fermentation these compounds decreased by more than 90% in total. OGS diminished by more than 90% in the lupin seeds. The PC content of the three seed species subjected to these processes increased their absorbance value, probably due to the enzymatic action of a fungal tannase. These results suggested that the L. mutabilis, L. campestris and G. max fermentation with R. oligosporus is an efficient method for diminishing antinutritional factors and for obtaining a product with optimal nutritional value. Copyright © 2007 Society of Chemical Industry     

芝麻酱、花生酱酸值和过氧化值测定中的油脂提取方法

为解决现有芝麻酱和花生酱酸值、过氧化值测定中油脂提取方法存在的提取时间长、效率低等问题,通过在样品中加入丙酮水溶液,使蛋白质发生变性、絮凝,从而破坏芝麻酱、花生酱稳定的溶胶体系,使体系快速分层,油脂快速游离出来,从而快速高效地提取油脂。具体方法：按料液比1∶1向芝麻酱、花生酱样品中加入丙酮水(体积比1∶1)溶液,搅拌混匀后加入3倍样品体积的石油醚,振摇提取,静置约30 min分层,将上层有机相经无水硫酸钠过滤,于40℃脱除溶剂,获得油脂。通过加入丙酮水溶液大大缩短了油脂提取时间,效率极大提高,并且降低了提取时间长带来的油脂氧化风险,提高了结果准确度。通过精密度、准确度方法学考察,证明该方法准确可靠。综上,该方法可以用于芝麻酱、花生酱产品酸值、过氧化值测定中油脂的提取。     

氮磷钾肥施用对油菜产量及养分吸收利用的影响

为油菜科学施肥提供依据,利用当前推广的6个油菜品种秦优11号（QY11）、中农油2008（ZNY2008）、中油杂11号（ZYZ11）、湘油17（XY17）、浙油601（ZY601）和沪油杂1号（HYZ1）,在大田试验条件下研究了氮、磷、钾肥施用对油菜产量及氮、磷、钾素吸收利用的影响,并比较了不同品种对施肥响应的差异。结果显示,相同施肥处理下不同品种籽粒产量差异显著,不施N（PKB）、不施P（NKB）、不施K（NPB）及NPK全施（NPKB）处理下品种间最大差异分别为385、244、759和720kg/hm2,变异系数分别为18．1％、25．5％、16．4％和11．0％。氮、磷、钾施用可显著提高各品种产量和相应养分积累量,NPKB处理相比PKB、NKB及NPB处理分别增产1101～2012、1681～2459和293～1567kg/hm2,N、P、K积累量分别增加63．0～113．2、17．2～23．8和94．1～166．3kg/hm2。不同品种氮、磷、钾肥利用率也存在显著差异。同一品种对氮、磷、钾的响应一致,其中秦优11号对氮、磷、钾肥施用的敏感程度均大于其它品种,湘油17耐低氮、低磷和低钾的能力均高于其它品种。     

Duplex real-time PCR method for the detection of sesame (Sesamum indicum) and flaxseed (Linum usitatissimum) DNA in processed food products

The development of a duplex real-time polymerase chain reaction (PCR) method allowing the simultaneous detection of sesame and flaxseed DNA in commercial food products is described. This duplex real-time PCR technique is based in the design of sesame- and flaxseed-specific primers based on the ITS1 region and two TaqMan fluorescent probes. The method was positive for sesame and flaxseed, and showed no cross-reactivity for all other heterologous plant and animal species tested. Sesame and flaxseed could be detected in a series of model samples with defined raw and heat-treated sesame in flaxseed, and flaxseed in sesame, respectively, with detection limits of 1.3 mg kg^?1 for sesame and 1.4 mg kg^?1 for flaxseed. The applicability of the assay for determining sesame and flaxseed in different food matrices was investigated by analysing a total of 238 commercial foodstuffs. This PCR method is useful for highly selective and sensitive detection of traces of sesame and flaxseed in commercial food products.     

The MAGi RNA extraction method: a highly efficient and simple procedure for fresh and dry plant tissues

BACKGROUND: Samples from different plant species, different organs or tissues at different times of the year, usually show great differences in their cell compositions, pH, and the endogenous RNase activities, decreasing the RNA yield and quality. RESULTS: In this study we describe a reagent and a simple total RNA isolation method for plant organs, tissues and dry seeds. The RNA extraction reagent (MAGi) is non‐toxic and can be stored at room temperature for several months to years. The principle of the total RNA extraction is that tissues are lysed in extraction solution with the aid of mortar homogenization–maceration, and cellular proteins, polysaccharides and DNA are removed from the RNA. We tested the reported method on more than 16 different types of plant seed and 15 different tissues and organs of pepper. CONCLUSION: The RNA extraction procedure reported in the present study greatly reduces the time required to isolate dry seed total RNA and other tissues by more than half as compared with the previously reported methods. The range of typical RNA yield and quality represents a significant improvement over existing protocols. The quality is high enough to be considered as suitable method for RT‐PCR, cDNA library construction and microarray gene expression studies. Copyright © 2008 Society of Chemical Industry     

油菜不同器官高质量总蛋白提取方法和双向电泳体系的优化

以显性核不育油菜Shaan—GMS不育株和可育株的花序和叶片为材料,采用TCA（三氯醋酸）-丙酮法提取油菜花蕾、花药及叶片总蛋白,对总蛋白提取方法、IPG（固定pH梯度）胶条种类、聚焦程序、凝胶浓度、上样量等环节进行了优化。结果表明,采用理想的蛋白质裂解液（7mol／L尿素,2mol／L硫脲,4％CHAPS（3-[3-（胆酰氨丙基）二甲氨基]丙磺酸内盐）,65mmol二硫苏糖醇,0．5％IPG缓冲液pH3～10或pH4～7,全蛋白酶抑制片剂（片／10mL））裂解蛋白,以150μg上样,按IEF程序Ⅱ（30V,12h;200V,1h;500V,1h;1000V,0．5h;10000V,2h;10000V,8h）进行等电聚焦,10％SDS—PAGE胶浓度进行第二向电泳,银染方法检测,可得到背景清晰、分辨率较高的油菜花蕾、花药及叶片蛋白质电泳图谱。高质量油菜蛋白的提取方法和蛋白质分离双向电泳体系的优化可为油菜蛋白质组学研究奠定技术基础。     

Optimization of extraction and isolation for 11S and 7S globulins of soybean seed storage protein

A new method was developed for extraction and isolation of 7S and 11S fractions from soybean seed, based on methods of Nagano et al., Thanh and Shibasaki [Nagano, T., Hirotsuka, M., & Mori, H. (1992). Dynamic viscoelastic study on the gelation of 7S globulin from soybeans. Journal of Agricultural and food chemistry 40, 941–944 and Thanh, V. H., & Shibasaki, K. (1976). Major proteins of soybean seeds. A straightforward fraction and their characterization. Journal of Agricultural and Food Chemistry 24, 1117–1121]. Optimization of the extraction and isolation of 11S and 7S globulins from soybean seed was investigated under various conditions by the Kjeldahl method and SDS-PAGE. The optimal conditions were as follows: 0.03–0.06 M Tris–HCl buffer (pH 8.5) containing 0.01 M sodium bisulfite as extract solution, extraction twice at 45 °C for 1 h, and with a 1:15 ratio (w/v) of flour:Tris–HCl. The 11S fraction was precipitated at pH 6.4, and the supernatant, after centrifugation, was adjusted to pH 5.5 to remove the insoluble intermediate fraction by further centrifugation. The supernatant obtained was then adjusted to pH 4.8 to afford the 7S fraction as a precipitate by centrifugation. With the improvements, the protein contents and purities of the isolated 11S and 7S fractions were significantly increased. The contents of all subunits of the isolated 11S and 7S fraction were markedly higher than those by Thanh and Shibasaki method, while the contents of α, β and B₃ were also significantly higher than those by Nagano et al. method.     

浓香型白酒酒醅中总RNA提取方法评价

为探寻我国特有的复杂微生态环境——浓香型酒醅样品总RNA提取方法,本实验建立了一种改良的十二烷基硫酸钠(sodium dodecyl sulfate,SDS)-苯酚法,并从提取成本及耗时、RNA质量浓度和纯度、电泳结果、反转录效果及高通量测序分析5 个方面,比较其与试剂盒法、Trizol法、十六烷基三甲基溴化铵(hex...