排除法PCR加变性梯度凝胶电泳鉴别未知基因

通过鉴别未知的cry4亚组基因来确定排除法PCR加变性梯度凝胶电泳新方法。方法 :应用排除法PCR的组基因和亚组基因引物扩增杀蚊毒素基因 ,cry4。这些引物是根据已知的cyr4基因的共有或特有DNA片段来设计的。组基因引物扩增产物被用于变性梯度凝胶电泳。平行变性梯度凝胶是由 8%的聚丙烯酰胺加上 2 0 %到 80 %的变性剂组成。结果 :已知的和未知的cry4亚组基因被组基因引物扩增的产物在变性梯度凝胶电泳被分离开。尽管它们之间仅有两个碱基对不同 (T在位置 2 2 4和G在位置 394)。应用组基因和亚组基因引物扩增 ,新发现的基因可被分类到次亚组基因水平。从 5个苏云金芽孢杆菌亚菌中发现三个未发表的cry4亚组基因。结论 :排除法PCR加变性梯度凝胶电泳是一个高度敏感、特异性和可靠性强的新方法 ,可用于鉴别各种未知的亚组基因。

Complete mitochondrial genome of the Five-dot Sergeant Parathyma sulpitia(Nymphalidae:Limenitidinae) and its phylogenetic implications

The complete mitochondrial genome of the Parathyma sulpitia （Lepidoptera, Nymphalidae, Limenitidinae） was determined. The entire mitochondrial DNA （mtDNA） molecule was 15 268 bp in size. Its gene content and organization were the same as those of other lepidopteran species, except for the presence of the 121 bp long intergenic spacer between trnSI（AGN）and trnE. The 13 protein-coding genes （PCGs） started with the typical ATN codon, with the exception of the coxl gene that used CGA as its initial codon. In addition, all protein-coding genes terminated at the common stop codon TAA, except the nad4 gene which used a single T as its terminating codon. All 22 tRNA genes possessed the typical clover leaf secondary structure except for trnSI（AGN）, which had a simple loop with the absence of the DHU stem. Excluding the A＋T-rich region, the mtDNA genome of P. sulpitia harbored 11 intergenic spacers, the longest of which was 121 bp long with the highest A＋T content （100%）, located between trnSI（AGN） and trnE. As in other lepidopteran species, there was an 18-bp poly-T stretch at the 3＇-end of the A＋T-rich region, and there were a few short microsatellite-like repeat regions without conspicuous macro-repeats in the A＋T-rich region. The phylogenetic analyses of the published complete mt genomes from nine Nymphalidae species were conducted using the concatenated sequences of 13 PCGs with maximum likelihood and Bayesian inference methods. The results indicated that Limenitidinae was a sister to the Heliconiinae among the main Nymphalidae lineages in this study, strongly supporting the results of previous molecular data, while contradicting speculations based on morphological characters.

表现为痉挛性截瘫的肾上腺脑白质营养不良一例

1病例报告患者男,47岁。主因“进行性步态异常伴排尿异常17年”于2019年6月入院。17年前(30岁)无明显诱因出现双下肢僵硬,间断有“抽筋感”,不能快速跑步,尿急。32岁时出现步态不稳,尿急明显,偶伴大便失禁,就诊当地医院,诊断为“运动神经元病可能”,给予神经营养治疗(具体不详)及巴氯芬40~50 mg/d,自觉下肢僵硬感稍好转,排便异常仍加重。37岁时患者不能独立站立,需持杖行走,自觉下肢发凉、少汗,尿不尽感明显,便秘,平均4~5 d排便1次。40岁时,患者出现性功能减退。近2个月,患者出现平卧起身困难。24年前患者曾发生颈部外伤,未留后遗症。否认家族中有类似病及遗传病史。入院查体:意识清楚,言语流利,全身皮肤未见色素沉着,心脏、双肺、腹部检查未见异常,高级神经活动正常,脑神经检查未见明显异常,四肢肌容积正常,双上肢肌张力正常,双下肢肌张力明显增高,双上肢肌力5级,双下肢肌力4级,双上肢指鼻试验稳准,双下肢跟膝胫试验不能配合,闭目难立征阳性,全身痛温觉未见明显异常,双上肢腱反射(++),双下肢腱反射(++++),双侧踝阵挛阳性,双侧Hoffmann征及Babinski征阳性,脑膜刺激征阴性。实验室检查:血、尿、便常规检查结果正常,肝肾功能、电解质、血糖正常,免疫全套、肿瘤标记物、甲状腺功能、抗中性粒细胞胞浆抗体、风湿及类风湿因子检测结果均正常,凝血功能、D-二聚体、淋巴细胞亚群均检测结果正常。脑脊液检查:压力180 mmH 2O(1 mmH 2O=9.8 Pa),常规及生化检测结果正常,IgG合成率正常,革兰染色、抗酸染色及墨汁染色、病毒学抗体均阴性,脑脊液寡克隆区带阴性。头MRI检查结果未见明显异常。颈胸椎MRI检查:颈椎退行性变,颈髓稍变细,胸髓均匀变细(图1)。泌尿系统超声检查:双肾、前列腺未见异常,残余尿44 mL。腹部超声检查:肝脏、胆囊、胰腺、脾脏及双肾未见异常。肌电图检查:双侧正中神经、尺神经、腓总神经、胫神经传导速度及波幅均正常,下肢针极肌电图正常。双下肢交感神经传导通路异常,心率变异图正常,体感诱发电位显示颈7至双顶皮层、胸12以上至双顶叶皮层深感觉传导潜伏期均延长。血极长链脂肪酸C2236.8 nmol/L(正常参考值,下同:<93.6 nmol/L),C2459.4 nmol/L(<91.4 nmol/L),C262.07 nmol/L(<1.3 nmol/L),C24/C221.61(<1.39),C26/C220.56(<0.23)。基因筛查提示存在ATP结合超家族D亚组膜1(ATP binding cassette subfamily D member 1,ABCD1)基因的第5号外显子cDNA1415位点AG缺失(图2)。后续随访显示,患者病情相对稳定,主要给予康复锻炼及对症治疗以改善生活质量。

Effect of ABCB1 C3435T Polymorphism on Clinical Outcomes in Kenyan HIV Patients on Lopinavir-Based Regimens

ATP Binding Cassette sub-family B member 1 （ABCB1） affects disposition of many drugs and thus affects the pharmacokinetics of drugs and ultimately treatment response. Polymorphisms of ABCB 1 especially ABCB 1 C3435T polymorphism may thus affect pharmacokinetics of antiretroviral drugs and hence CD4 treatment response and other clinical outcomes of HIV patients. Methods： The study design was a historical cohort study and entailed collection of patient data. PureLink genomic DNA extraction mini kit was used for the extraction and purification of genomic DNA. TaqMan drug genotyping assay and protocol was used in the DNA amplification and genotyping. Data analysis was done using STATA software version 10. Results： Study participants with the CT genotype had lower creatinine levels after 6 months on lopinavir-based regimens compared with those with the CC genotype （p = 0.001）. In addition, the study participants with the CT genotype had consistently higher CD4 cell counts compared with those with the CC genotype from the time of ART switch but this was not statistically significant. However, there was no significant association between the ABCB 1 C3435T genotypes and haemoglobin and ALT levels. Conclusion： There was a significant association between ABCB1 C3435T polymorphism and creatinine levels 6 months after therapy on lopinavir-based regimens.

Study on polymorphism ofArtemisia capillaris by using RAPD for Terengganu population in Malaysia

Artemisia capillaris is a herbaceous aromatic and therapeutic plant. The genetic variability among individuals of Artemisia capillaris from state of Terengganu, Malaysia was examined by using the random amplified polymorphic DNA （RAPD） technique to assess the polymorphism at the species level, The samples from differences regional in Terengganu State. The genomic DNA was extracted from the samples leaves using Sarkosyl method. The results produced by the machine showed clear RAPD banding pattern. Fifty-seven oligonucleotide primers were screened and five primers were selected （OPA 04, OPA 09, OPA 16, OPA 17 and OPA 18） to amplify DNA from five samples of Artemisia capillaris from State of Terengganu, Malaysia. A total of 135 RAPD fragments （RAPDs） with all polymorphic fragments （100%） with size ranging from 250--3000 bp were scored from the population. Genetic distance for samples ranges from 0.0000 to 0.320000. For similarity index samples ranges from 0.0000 to 0.7547.

Isolation, identification and genomic characterization of the Asian lineage Zika virus imported to China

Dear Editor,Zika virus（ZIKV）used to be an unknown mosquito-borne flavivirus,and maintained its limited sylvatic circulation in a few African and Asian countries（Enfissi et al.,2016）.Based on available clinical data,the symptoms in human infections with ZIKV are supposed to be similar to other arbovirus infections such as dengue,and characterized by fever,skin rashes,conjunctivitis,muscle and joint pain,malaise,and headache（Duffy et al.,2009）.