应用PCR-RFLP和芯片生物分析系统鉴别河豚鱼品种

应用聚合酶链式反应-限制性片段长度多态分析和芯片生物分析系统建立5种河豚的分子生物学品种鉴定新方法。首先提取鱼的DNA进行细胞色素b基因的PCR扩增,然后用DdeⅠ、HaeⅢ和NlaⅢ三种限制性内切酶进行酶切,在Agilent DNA1000芯片上对酶切片段进行分离。该方法可成功区分5个河豚鱼品种,是一种快速、简便、有效的鱼类品种鉴定分析手段。     

13 种河豚鱼CO I基因部分DNA序列分析及在物种鉴定中的应用

应用CO I基因聚合酶链式反应（polymerase chain reaction,PCR）扩增通用引物,对福建省和海南省搜集的3?属13?种河豚鱼样品与9?个未知种名的河豚鱼样品的CO?I基因靶序列片段进行PCR扩增和测序,13?个已知物种样品的DNA序列提交基因库（GenBank）,取得相应的登录号。各物种CO I基因靶序列长度均为681 bp。应用DNAMAN V6软件对样品的DNA序列进行同源性比对分析,建立样品间系统关系同源树。基于CO I基因序列,供试13 个样品被划分为4?个类群组。群间的同源率为84%,群内同源率为90%～100%。根据序列同源性分析结果,9?个未知种名的样品被归类到2?个类群组中,判定这9?个样品为东方鲀属或腹刺鲀属,其中1?个样品（HNW2）为月腹刺鲀,4?个样品（HNW3、HNW4、FJW2、FJW5）为棕斑腹刺鲀,1?个样品（HNW1）为暗鳍腹刺鲀;3?个样品（FJW1、FJW3、FJW4）为横纹东方鲀。探讨基于DNA条形码技术的CO I基因靶序列片段在河豚鱼种属鉴别中应用的可能性。     

Identification of Toxin and Fish Species in Cooked Fish Liver Implicated in Food Poisoning

ABSTRACT: The cooked fish liver retained by the victims was assayed for toxicity and mitochondrial DNA. Meanwhile, 8 live specimens of puffer Takifugu niphobles were also assayed. The toxicity of cooked fish liver was 280 ± 20 mouse units per gram (MU/g). All specimens of T. niphobles showed high toxicity (more than 850 MU/g) in the liver. The toxin from cooked fish liver and liver of T. niphobles was identified to be tetrodotoxin. The cooked fish liver and fresh liver of T. niphobles showed the same sequence genotype and the same single restriction site for Bsa I. Therefore, the species of cooked fish liver was suggested as T. niphobles.     

河豚鱼Cyt b基因部分DNA序列分析与应用

建立河豚鱼物种DNA鉴别技术,根据GenBank公布的河豚鱼细胞色素b基因序列,应用软件Primer Premier5.00版设计上游引物HT1-F与下游HT1-R,对3属9种福建省搜集的常见的河豚鱼与2种未知物种名称且外观无法进行形态学判断的河豚鱼样品的细胞色素b基因序列中的部分片段进行PCR扩增,PCR产物通过琼脂糖凝胶电泳确证、纯化后,进行DNA碱基序列测定,序列长度均为423bp。应用DNA MANN软件进行样品间DNA序列同源性比对分析,建立样品间系统关系树状图,供试11个样品被划分为3个类群,第Ⅰ组与第Ⅱ组之间的同源率为89%,这2组与第Ⅲ组之间的同源率为85%。根据序列同源性分析结果,可推测2个未知种名的样品为腹刺鲀属或东方鲀属。     

Capillary Zone Electrophoresis of Fish Muscle Sarcoplasmic Proteins

A capillary zone electrophoresis (CZE) method for rapid separation of fish muscle sarcoplasmic proteins (SAJR) was developed using a highionic-strength phosphate buffer and an untreated, fused silica capillary (20 μm i.d. X 20 cm). Changes in pH between 6.6-7.5 resulted in differences in mobility, separation factors, and number of SAR moieties. An optimal SAR profile was obtained at pH 7.4. When used to monitor frozen storage changes in SAR from cod (Atlantic, Ling, Pacific) and pollock fillets, the procedure indicated both species differences and low-and high- molecular-weight frozen storage changes. CZE is an effective screening/quantification alternative to conventional electrophoresis and/ or HPLC because it is fast (<10 min), simple, sensitive, precise, and readily automated.     

河豚鱼16S rRNA基因部分DNA序列分析及应用

应用16S rRNA基因聚合酶链式反应（polymerase chain reaction,PCR）扩增通用引物,对3 属13 种福建省搜集的河豚鱼样品与9 个未知物种的河豚鱼样品的16S rRNA基因序列中的部分片段进行PCR扩增与脱氧核糖核酸（deoxyribonucleic acid,DNA）碱基序列测定,各物种序列长度在611～614 bp之间。应用DNAMAN V6软件进行样品间DNA序列同源性比对分析,建立样品间系统关系同源树。供试13 个样品被划分为4 个类群组,群间的同源率为87%,群内同源率为94%～100%。根据序列同源性分析结果,9 个未知种名的样品被归类到3 个类群组中,推测9 个未知种名的样品为东方鲀属或腹刺鲀属。探讨了16S rRNA基因部分DNA序列测试及同源性分析技术在河豚鱼种属鉴别中应用的可能性。     

骨素及骨素酶解液中蛋白质及肽的电泳分离

为了分离纯化骨素及其酶解产物中的蛋白质及肽,并比较骨素蛋白与骨蛋白和肌肉蛋白的差异,本文以骨蛋白和肌肉蛋白为参照,采用十二烷基磺酸钠凝胶电泳法(SDS-PAGE)对骨素蛋白及其酶解产物中的肽进行分离。利用不同浓度的分离胶(5%、7.5%、10%、15%)对骨素蛋白、骨蛋白和肌肉蛋白样品进行分离,结果发现,各种浓度的分离胶均可以分离骨蛋白和肌肉蛋白,但是不能分离骨素蛋白。对骨素蛋白酶解液采用N-三(羟甲基)甲基甘氨酸凝胶电泳法分离,能有效分离骨素酶解液中的肽。对SDS-PAGE电泳分离时骨蛋白的迁移比和分离胶的相关性分析发现,骨蛋白的迁移比与分离胶浓度间呈指数相关,且具有极显著的相关性(R2≥0.996,p0.01)。利用该相关性并结合蛋白质的分子量与迁移比的相关性方程,可以通过蛋白质的分子量选择合理的分离胶浓度,或者通过迁移比预测蛋白质的分子量。     

Biolog微生物鉴定系统在菌种鉴定中的应用

Biolog微生物鉴定系统由浊读仪、读数仪、软件、数据库、微孔鉴定板组成，与计算机联网。对结果的鉴定需要考虑可能性、相似性、位距3个参数；通过SIM，DIST和PROB值与数据库中的ID比较，确定被鉴定的微生物的属种。     

两种织纹螺肌肉蛋白的电泳鉴别

利用聚丙烯酰胺凝胶电泳(SDS-PAGE)对有毒的光织纹螺(Nassariusrutilans)和无毒的西格织纹螺(Nassariussiquinjorensis)的肌肉蛋白做鉴别分析。实验采用四种不同抽提液对肌浆蛋白和肌原纤维蛋白进行提取。肌肉蛋白经电泳后以考马氏亮蓝染色并对蛋白图谱作对比分析。根据蛋白条带的数量、迁移率及浓度等特点,比较得出两种织纹螺的差异条带。综合比较得出,前者的肌肉蛋白特异条带为34.9kDa、32.7kDa,后者的特异条带为36.0kDa。结果表明,以SDS-PAGE电泳法可实现对该两种织纹螺的初步鉴别。     

A Rapid Method for the Recovery,Quantification and Electrophoretic Analysis of Proteins from Beer

A previously‐developed method for protein recovery from wine has been applied to beer and beer foam samples. The method involves the complexation of proteins with dodecyl sulfate (added as sodium salts) and subsequently the insolubilization of the protein‐detergent complexes by addition of potassium ions (added as KCl). The protocol allows preparation of proteins from a few hundred microliters of beverage in a few minutes. The precipitated proteins are free from interfering materials and are directly utilizable for quantitative and electrophoretic assays.     

Identification of Red Snapper (Lutjanus campechanus) using Electrophoretic Techniques

Isoelectric focusing (IEF), sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and two-dimensional (2-D) gel electrophoresis were employed to produce protein profiles for species identification of red snapper and 11 other fish species. Comparing the distinctive patterns of water-soluble sarcoplasmic proteins for each species on IEF and SDS gels, red snapper could be identified. IEF gels of ampholyte mixture of 20% pH 3–10 and 80% pH 4–6.5 resolved better than gels with ampholyte ranging from pH 3–10, 4–6.5, 6–8, or 5–6 for species identification. The 10 and 12.5% SDS-PAGE gels produced more distinctive protein profiles for identification than 7.5 and 15% gels. Thus, these techniques could be applied to identify fish species.     

大豆致敏蛋白的识别

我国是大豆的种植大国，但对大豆致敏蛋白的识别却一直没有进行过，本文就是利用大豆过敏患者的血清识别不同品种大豆致敏蛋白的组成，除已被发现的7S(β-伴大豆球蛋白)外，还识别到一种分子量85．3kDa的强致敏蛋白，同时发现，品种间的差异蛋白都有可能成为新的致敏蛋白。     

4种养殖河豚鱼肌肉营养成分分析与评价

分析4种河豚鱼肌肉中氨基酸,脂肪酸和矿物质元素等营养成分,并评价其营养价值,为市场发展河豚鱼产业提供数据支持。分别采用干燥法、灰化法、凯氏定氮法、气相色谱法、氨基酸自动分析仪、电感耦合等离子体质谱仪测定4种河豚鱼肌肉中的常规营养成分、氨基酸、脂肪酸和矿物质的含量。结果表明,4种河豚鱼肌肉中的蛋白质含量丰富,脂肪含量较少,蛋白质含量和脂肪含量最高的分别是菊黄东方鲀和红鳍东方鲀。其中,双斑东方鲀的必需氨基酸与总氨基酸的比值(essential amino acid/total amino acid,EAA/TAA)、必需氨基酸与非必需氨基酸的比值(essential amino acid/non-essential amino acid,EAA/NEAA)和必需氨基酸评分(essential aamino acid index,EAAI)最高,且符合联合国粮农组织/世界卫生组织评分(Food and Agriculture Organization/World Health Organization,FAO/WHO)标准,红鳍东方鲀的总饱和脂肪酸(total saturated fatty...     

基于DNA条形码技术鉴别有毒鹅膏菌属物种

收集27个鹅膏菌属物种共38份样本,提取样品基因组DNA,应用通用引物扩增其内转录间隔区(internal transcribed spacer,ITS)、核糖体大亚基(large ribosomal subunit,LSU)、RNA聚合酶的第二大亚基(the second largest subunit of RNA...     

DNA条形码技术对鱼子酱物种溯源及鉴定

为探讨DNA条形码技术在鱼子酱物种鉴定中的适用性,利用细胞色素b（Cytochrome b,Cyt b）和细胞色素氧化酶I亚单位I（Cytochrome Oxidase I,COI）作为DNA条形码对鱼子酱样品进行DNA提取、聚合酶链式反应（Polymerase Chain Reaction,PCR）、测序、利用NCBI网站和BOLD鉴定系统进行基因比较分析,构建系统发育树,鉴定鱼子酱物种,对我国鱼子酱产品物种标签符合性情况进行检查。购买的40份样品,一致性鲟鱼物种基因序列相似性均在99%以上,涉及5个鲟鱼种,其中杂交种占比75%、西伯利亚鲟、施氏鲟、欧鳇、俄罗斯鲟占25%。说明Cyt b、COI作为DNA条形码可以对鱼子酱进行物种鉴定,检测的鱼子酱产品均为鲟鱼子酱,无造假,但是45%产品标签物种替代或物种标识不清。加强对产品物种标识重视及鉴定技术的开发,有助于我国鱼子酱对外贸易发展,保障我国鲟鱼产业可持续性健康发展。     

食品中肉类成分种属鉴别技术研究进展

肉与肉制品的掺杂、掺假是食品质量控制面临的重要挑战。食品中肉类成分的鉴别与分析技术已逐步形成了分别以蛋白质检测和以核酸检测为基础的方法体系,其鉴别精度可达属与亚属水平,检测灵敏度也达到了纳克级。近年来,食品中肉类成分的定量检测与溯源又成为了本领域中的新研究热点。本文综述食品中肉类成分种属鉴别技术的研究进展,并重点分析应用荧光定量PCR对食品中肉类成分进行定量分析的现状与前景。     

Identification of Meat from Game and Domestic Species

Isoelectric focusing combined with specific enzyme staining for creatine kinase was used to characterize banding patterns in meat from pronghorn, mule deer, white-tailed deer, sheep, moose, pork, bison, elk, caribou, red deer, beef and goat. Processed and cooked pork was differentiated from all species and beef and elk were separated from pronghorn. It was not possible to differentiate beef from elk, or prong-horn from sheep. The inability to separate some game from some domestic species and the lability of the staining proteins after heating above 67°C limits the application of this technique.     

Electrophoretic Identification of Fish Species Used in Surimi Products

Both sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and urea gel isoelectric focusing (IEF) were used to identify species-specific protein bands of raw and cooked fish and surimi samples from Alaska pollock (Theragra chalcogramma) and red hake (Urophycis chuss). In raw samples, species-specific bands were found in the water extracts, while in cooked samples 1% SDS and 8M urea extracts were more effective for species identification in both fish and surimi.     

蛋白质组学技术在鉴定啤酒浑浊蛋白中的初步研究

啤酒中包含多种大麦蛋白,制麦和酿造过程中受到化学方式或酶的修饰,影响最终的啤酒浑浊稳定性。从啤酒中分离出的浑浊活性蛋白主要来自大麦储藏蛋白或大麦醇溶蛋白（Hordeum vulgare L.）,这些蛋白有富含脯氨酸,由许多不同分子量的片段组成。尽管对啤酒浑浊的研究时间较长,但是对于其特性方面的有待进一步确认。     

冷藏温度对河豚鱼微生物及品质特性的影响

研究冷藏温度对河豚鱼微生物及品质特性的影响。将河豚鱼宰杀、清理、分块后,放置于(0±1)、(4±1)、(8±1)℃冰箱中冷藏。对鱼肉中几种优势菌种(肠杆菌、假单胞菌及乳酸菌),鱼肉质构、挥发性盐基氮、pH值、三甲胺、持水力等指标进行测定,评价河豚鱼冷藏过程中的品质变化。结果表明：(0±1)℃可贮藏6d左右,能够一定程度地抑制鱼肉中几种特定腐败菌的生长,减慢TVB-N值、pH值和TMA值的上升,延缓持水力的下降,并降低河豚鱼肉的硬度、弹性和回复性等质构特性的变化,有效延长河豚鱼的货架期,维持良好的鲜度。