猪圆环病毒Ⅰ型ORF1基因的克隆及其在大肠杆菌中的表达

根据GenBank中猪圆环病毒Ⅰ型(PCV1)ORF1基因序列,设计合成了1对引物,对PCV1 ORF1基因进行PCR扩增,将扩增出的ORF1基因(939 bp)克隆入pMD18-T载体,将筛选获得的重组质粒命名为pMD-ORF1。测序分析表明,克隆的ORF1与广东分离株DQ659154的核苷酸序列同源性高达99.6%,推导的氨基酸序列同源性为99.4%。将ORF1双酶切产物插入原核表达载体pET-41a,得到的重组子命名为pET-ORF1。用IPTG进行诱导表达,收集菌液进行SDS-PAGE和Western-blot分析,结果表明,PCV1ORF1在pET-41a中获得了高效融合表达,其表达蛋白的分子质量约为67.6 ku。     

Phenolphthalein False‐Positive Reactions from Legume Root Nodules

Presumptive tests for blood play a critical role in the examination of physical evidence and in the determination of subsequent analysis. The catalytic power of hemoglobin allows colorimetric reactions employing phenolphthalein (Kastle‐Meyer test) to indicate “whether” blood is present. Consequently, DNA profiles extracted from phenolphthalein‐positive stains are presumed to be from blood on the evidentiary item and can lead to the identification of “whose” blood is present. Crushed nodules from a variety of legumes yielded phenolphthalein false‐positive reactions that were indistinguishable from true bloodstains both in color quality and in developmental time frame. Clothing and other materials stained by nodules also yielded phenolphthalein false‐positive reactivity for several years after nodule exposure. Nodules from leguminous plants contain a protein (leghemoglobin) which is structurally and functionally similar to hemoglobin. Testing of purified leghemoglobin confirmed this protein as a source of phenolphthalein reactivity. A scenario is presented showing how the presence of leghemoglobin from nodule staining can mislead investigators.     

Development of a Tetraplex PCR Assay for CYP2D6 Genotyping in Degraded DNA Samples

CYP2D6 polymorphism analysis is gaining increasing interest in forensic pharmacogenetics. Nevertheless, DNA recovered from forensic samples could be of poor quality and not suitable for long polymerase chain reaction required to type CYP2D6 gene prior to SNaPshot minisequencing analysis performed to define alleles with different enzymatic activity. We developed and validated following the guidelines of the Scientific Working Group on DNA Analysis Methods a tetraplex PCR yielding four amplicons of 597, 803, 1142, and 1659 bp encompassing the entire CYP2D6 gene to analyze eleven SNP positions by SNaPshot minisequencing. Concordance, sensitivity, and specificity were assessed. The method, applied to thirty‐two forensic samples failed to amplify with long PCR, allowed the amplification of CYP2D6 gene in 62.5% of degraded samples. The new tetraplex PCR appears a suitable method for CYP2D6 analysis in forensic pharmacogenetics.     

Legislative genesis and judicial death of a directive: The European Court of Justice invalidated the data retention directive (2006/24/EC) thereby creating a sustained period of legal uncertainty about the validity of national laws which enacted it

The Grand Chamber has ruled that the data retention directive was invalid ex tunc since it seriously interfered with the fundamental rights to respect for private life and protection of personal data and exceeded the limits of the principle of proportionality which are provided for in the Charter. The scope and temporal effects of this ruling should be clarified, especially its legal impacts on national laws of Member States which enacted the directive. In addition, the findings of the Grand Chamber on geographical safeguards have far-reaching implications on the retention and storage of personal data in the EU.     

小鼠脑外伤后Apelin-13对自噬相关蛋白表达的影响

目的观察Apelin-13对小鼠脑外伤（Traumaticbraininjury,TBI）自噬相关蛋白的影响,并初步探讨其对小鼠脑外伤后自噬相关蛋白调节的作用机制。方法取健康雄性CD-1小鼠,随机分为假手术（sham）组、生理盐水（sa-line）组、Apelin-13组。Apelin-13组和saline组在建立脑外伤模型前10min,分别注射Apelin-13（0．3μg／g）或等量生理盐水于侧脑室,然后利用Western-Blot检测脑外伤后24h、48h组大脑皮质自噬相关蛋白LC3、P62、Beclin-1和Bcl-2等的表达情况。结果与sham组相比,在脑外伤后24h、48h,saline组自噬相关蛋白LC3-Ⅱ／LC3-Ⅰ比值,Beclin-1的蛋白表达均被显著上调（P〈0．01）;同时Bcl-2、P62的蛋白表达水平被显著下调。然而,与saline组相比,Apelin-13组可显著下调自噬相关蛋白LC3-Ⅱ／LC3-Ⅰ比值和Beclin-1的蛋白表达水平（P〈0．01）;同时显著上调Bcl-2和P62的表达水平;亦明显降低Beclin-1／Bcl-2的比值。结论Apelin-13可以明显抑制脑外伤后小鼠大脑皮质细胞的自噬活性,这一作用可能是通过调节Beclin-1、Bcl-2蛋白表达水平来实现的。     

PowerPlex 21与GoldeneyeTM 20A试剂盒的一致性验证及法医学应用

目的：确认PowerPlex 21试剂盒与GoldeneyeTM 20A试剂盒分型结果的一致性。方法应用两试剂盒对205名北京汉族无关个体血样DNA进行复合扩增,观察19个重叠STR基因座分型的一致性,并统计D1S1656的遗传多态性。结果所有19个重叠基因座分型相同,两个试剂盒的杂合基因座峰高比例差异无统计学意义（P〉0.05）。D1S1656杂合度为0.878,个人识别率为0.949,三联体非父排除率为0.751,二联体非父排除率为0.506,多态信息含量为0.810。结论 PowerPlex21试剂盒与GoldeneyeTM 20A试剂盒分型结果一致性好,引物设计合理;D1S1656多态性好,可用于人类遗传分析及法医学中的亲子鉴定和个人识别。     

国产NuHi-POP4与ABI POP-4在毛细管电泳分离效果的比较

目的比较国产基因分离胶NuHi-POP4与美国产商品ABI POP-4在法医鉴定应用中的分离性能。方法以国产基因分离胶NuHi-POP4和美国产的ABI POP-4为研究对象,分别考察其在毛细管电泳过程中对标准品LIZ500和实际样品的分离效果,并统计其电泳参数,包括电泳时间、电流、单碱基分辨率以及线性关系等,从而来比较两种胶的分离性能,评价其性能。结果 NuHi-POP4与ABI POP-4相比,不仅可以显著缩短内标和样品的分析时间,同时还表现出电流稳定、分辨率高、线性关系好等特点。结论国产NuHi-POP4与美国ABI POP-4相比,同样具有稳定、快速、高分辨等优势,结果准确可靠。     

美国“Z世代”大学生政治认同两极化及其原因

美国“Z世代”大学生的政治认同呈现明显两极化、主体向左的趋势。一方面,这种两极化与两党意识形态、政策立场保持高度一致;另一方面,两极化也显示出新的代际特征,即鲜明的道德色彩与强烈的对抗性。从社会心理学角度看,政治认同两极化的原因包括信念激情期的周期性再现、美国大学校园成为政治认同极化的孵化器、安全空间建构与微侵犯观念隔断大学不同政见者的沟通与交流、社交媒体强化“Z世代”同一阵营内部的政治认同等。“Z世代”大学生以“对抗共同敌人”的方式参与2020年美国总统选举并使投票率创下新高,这成为其政治认同极化公开表达的明证。这一态势引起了专家、学者等的警惕,重建大学校园政治生态成为美国社会的共识。大学生政治认同两极化也给未来的中美关系增添了不确定因素,社交媒体的运用将使“Z世代”大学生对华民意更具敌意,受害者心理将使美国舆论在人权等相关问题上对中国持更负面的评价,安全空间防线将使中国对外文化传播受到进一步干扰。“Z世代”大学生政治认同两极化趋势及大学校园政治生态将是评估美国政治未来走向的重要参考因素。     

犬Mnx1基因过表达载体的构建及其对脂肪间充质干细胞向胰岛β细胞分化的影响

本研究旨在构建犬Mnx1基因腺病毒过表达载体,并探究Mnx1基因过表达对犬脂肪间充质干细胞向胰岛β细胞方向分化的影响。本试验构建了腺病毒重组表达载体pAD-Mnx1,经293A细胞包装扩增后产生具有感染性的重组腺病毒,将其按感染复数(MOI)100感染犬ADSCs后,分别在第3、6、9、12和15天观察绿色荧光蛋白的表达情况、细胞形态变化和胰岛β细胞发育相关基因的表达情况。结果显示,ADSCs在感染后24 h出现绿色荧光,形态无显著变化;Mnx1基因表达可激发胰十二指肠同源盒1(Pdx1)基因、神经元素3(Ngn3)基因、肌腱膜纤维肉瘤癌基因同源物A(Mafa)基因、NK2同源盒2(Nkx2.2)基因、NK6同源盒1(Nkx6.1)基因、神经分化因子1(Neurod1)基因、成对合基因4(Pax4)和GATA结合蛋白4(Gata4)基因等内源性基因的表达。结果证实,腺病毒介导的过表达载体pAD-Mnx1可在犬ADSCs超表达Mnx1基因,从而诱导犬ADSCs向胰岛前体细胞分化,具有进一步诱导犬ADSCs形成胰岛β细胞的可能。