农产品中农药残留分析技术研究进展

农药残留的极性中毒事件和农产品进出口贸易中农药残留超标问题时有发生,农药残留分析是保证人民群众食品安全和进出口企业利益的重要手段,近年来农产品中农药残留分析技术有了快速的发展。本文主要在农药残留分析中的样品前处理和仪器检测2方面对近5年来的研究进展进行分析和讨论,主要探讨了前处理中的QuEChERS方法的应用、改进和发展,以及仪器检测手段中串联质谱的应用现状及高分辨质谱的应用前景,最后对农药残留分析这两方面的发展进行了展望。     

CRISPR/Cas9介导的ASGR1基因敲除猪制备

猪(Sus scrofa)内皮细胞的去唾液酸糖蛋白受体1(asialoglycoprotein receptor 1,ASGR1)是诱发异种肝移植后受体发生血小板减少症的主要诱因之一.本研究在α-1,3-半乳糖基转移酶基因(α-1,3-galactosyltransferase,GGTA1)敲除的五指山小型猪基础上,利用规律成簇间隔短回文重复(clustered regularly interspaced short palindromic repeats/Cas9,CRISPR/Cas9)结合体细胞核移植技术制备ASGR1基因敲除猪,针对猪ASGR1基因设计并构建了靶向表达载体单链导向RNAl (single guide RNA1,sgRNA1),将sgRNA1与增强绿色荧光蛋白(enhanced green fluorescent protein,EGFP)质粒共同电转染GGTA1基因敲除猪耳成纤维细胞,流式细胞仪富集筛选带绿色荧光的细胞后培养单细胞克隆.实验结果表明,PCR测序检测培养获得的41个单细胞克隆,37个克隆发生基因突变,ASGR1基因突变效率为90％,其中双等位基因敲除的细胞克隆30个,效率高达73％.选择4个ASGR1基因敲除类型不同的细胞为核供体进行核移植,将早期重构胚移植到4头受体猪,2头妊娠至终期,产仔猪6头,Western blot显示ASGR1基因敲除仔猪的肝脏中没有ASGR1的表达.对sgRNA1的13个潜在脱靶位点,分别设计引物进行PCR扩增和测序,结果显示没有脱靶现象.总之,本研究利用CRISPR/Cas9结合体细胞核移植技术快速高效地制备了GGTA1/ASGR1基因敲除五指山小型猪模型,有望解决异种肝移植后出现的血小板减少症,为临床过渡肝的应用研究提供了良好的研究材料.     

Abundance, polymorphism and genetic mapping of microsatellites in oilseed rape (Brassica napus L.)

The objective of the present study was to estimate the abundance and degree of polymorphism of simple sequence repeat (SSR) markers in rapeseed. By screening about 45000 clones of a small inserts library of rapeseed total DNA the abundances of GA/TC and CA/TG simple sequence repeats in the rapeseed genome were estimated to be approximately one repeat every 100 kb and 400 kb, respectively. After sequencing 13 positive clones, primer pairs could be designed for 11 microsatellite loci. Seven of these primer pairs produced reproducible amplification products in a set of 31 rapeseed genotypes, with one pair amplifying two independent products, giving a total of eight amplified loci. The different microsatellite loci displayed between one and three visible alleles. At four loci, additional null alleles were observed. With up to four alleles, polymorphic microsatellite markers show significantly higher allele numbers in rapeseed than restriction fragment length polymorphism (RFLP) markers. Four of the eight microsatellite markers could be mapped on four different linkage groups of an RFLP map of the rapeseed genome.     

A study of the disappearance of pesticides during composting using a gas chromatography-tandem mass spectrometry technique

A simple method for multi-residue analysis of pesticides by low-pressure gas chromatography-tandem mass spectrometry (LP-GC-MS-MS) has been validated in compost samples. The pesticide residues were extracted from the lyophilised samples with organic solvent by stirring. No sample clean-up was required prior to the analysis. The method was applied to determine the fate of two organochlorine pesticides (lindane and endosulfan) and two organophosphates (malathion and chlorpyrifos-methyl) during the composting process in the reactor of a pilot plant. Malathion, chlorpyrifos-methyl and lindane residues almost fully disappeared after 8 days of maturation in the reactor, while endosulfan residues were only partially degraded. Alongside this, a study of natural disappearance of the pesticides was carried out in the laboratory and a characteristic time profile was traced for each pesticide class.     

Analysis of differentially expressed proteins in the pollen of Malus spectabilis (crabapples) and Malus domestica (‘Gala’ apples) and their role in pollination

Our objective was to study the differentially expressed proteins (DEP) in various Malus spectabilis (crabapple) varieties (M. ‘Snowdrift’, M. ‘Hongling’ and M. ‘Hongjin’) in relation to Malus domestica (‘Gala’) and their role in pollination. Our method used a two-dimensional electrophoresis (2-DE) to analyse the differential proteins in the pollen of several crabapples. The 2-DE apples combined with the tandem mass spectrometry (MS-MS) and protein database retrieval helped us to identify the nature and function of DEPs in ‘Gala’ apples and crabapples. We identified 1195 proteins through 2-DE. Among these, six DEPs, namely chloroplast ferritin, Actin, Beta-fructofuranosidase, vacuolarH+-ATPase catalytic subunit, Full = Phosphoglucomutase, and Cytochrome b were identified by MS-MS. This study identified six DEPs among the pollen from the ‘Snowdrift’ crabapple, ‘Hongling’ crabapple, ‘Hongjin’ crabapple, and ‘Gala’ apples. The DEPs included metabolism related proteins, stress/regulatory proteins, and proteins involved in signal transduction.     

喹乙醇在鲫体内消除规律的研究

报道了鲫单剂量口灌50mg／kg喹乙醇在血浆、肌肉和肝胰脏中消除规律的研究。用乙腈-乙酸乙酯（1／1：V/V）提取组织中的喹乙醇,血浆中喹乙醇用3％的三氯乙酸沉淀蛋白,乙酸乙酯和磷酸盐缓冲液提取喹乙醇代谢物3-甲基喹嗯啉-2-羧酸,超高效液相色谱串联质谱法测定血浆及组织中喹乙醇及其代谢物的浓度。鲫单剂量口灌喹乙醇50mg／kg后,在血浆、肌肉和肝胰脏中的喹乙醇随着时间的延长呈现不同浓度的降解趋势。喹乙醇在血浆中的消除需要4d,在肌肉和肝胰脏中需要7d;喹乙醇的代谢物3-甲基喹嗯啉-2-羧酸在鲫体内消除比较缓慢,肌肉中需要15d,肝胰脏中需要20d。     

圆斑星鲽mtDNA控制区结构

对圆斑星鲽mtDNA控制区序列及鲽形目鱼类鲽科的条斑星鲽、大西洋黄盖鲽、马舌鲽，美洲拟庸鲽和鲆科的牙鲆以及鳎科的欧洲鳎、塞内加尔鳎、沙鳎控制区进行了比较分析。结果表明，圆斑星鲽的控制区序列可分为扩展终止相关序列区（ETAS）、中央保守区（CSB—A、B、C、D、E、F）、保守序列区（CSB1、CSB2、CSB3）和串联重复序列区（Tandem repeat region）。通过与其他脊椎动物线粒体控制区序列的比较，发现鲽形目的鲆、鲽类和鳎类与两栖纲的无尾类在CSB-3之后存在相似的串联重复序列。     

气相色谱法测定芒果中的苯醚甲环唑残留

通过考察4 种不同的样品前处理方法,建立了芒果中苯醚甲环唑残留量的气相色谱法快速分析方法。样 品经乙腈涡旋提取,通过PSA 和无水MgSO4高速离心净化,毛细管柱分离,气相色谱-电子捕获检测器（GC-ECD）检 测,外标法（基质加标）定量。结果表明院该方法在苯醚甲环唑添加量为2~200 滋g/L 范围内线性相关性良好,R2= 0.9991,最低检出限为0.005 mg/kg。样品在0.05、0.50、2.00 mg/kg 3个添加水平下的苯醚甲环唑回收率为90.5%~ 98.0%,相对标准偏差（RSD）为1.56%~3.40%。与另外3 种方法相比,该方法具有操作简便、处理时间短、检测成本低、 灵敏度高、定量准确等诸多优点,适合大批量样品的检测,已成功应用于实际样品的检测。     

Structure of genetic diversity among common bean (<Emphasis Type="Italic">Phaseolus vulgaris</Emphasis> L.) varieties of mesoamerican and andean origins using new developed microsatellite markers

A common bean genomic library was constructed using the ‘IAC-UNA’ variety enriched for (CT) and (GT) for microsatellite motifs. From 1,209 sequenced clones, 714 showed microsatellites distributed over 471 simple and 243 compound motifs. GA/CT and GT/CA were the most frequent motifs found among these sequences. A total of 123 microsatellites has been characterized. Out of these, 87 were polymorphic (73.7%), 33 monomorphic (26.8%), and 3 (2.4%) did not amplify at all. In a sample of 20 common bean materials selected from the Agronomic Institute Germplasm Bank, the number of alleles per locus varied 2–9, with an average of 2.82. The polymorphic information content (PIC) of each marker varied from 0.05 to 0.83, with a 0.45 average value. Cluster and principal coordinate analysis of the microsatellite data were consistent with the original assignment of the germplasm accessions into the Andean and Mesoamerican gene pools of common bean. Low polymorphism levels detected could be associated with the domestication process. These microsatellites could be a valuable resource for the bean community because of their use as new markers for genetic studies. Electronic supplementary material The online version of this article (doi: ) contains supplementary material, which is available to authorized users. An erratum to this article can be found at