MGB(Minor Groove Binder)复合探针二重实时定量PCR检测大肠杆菌O157

以大肠杆菌O157rfbE和stx2为待检靶基因,设计两对引物和两条MGB探针,rfbE和stx2探针5′端分别用FAM和VIC基团标记,3′端均用Taqman-MGB标记。建立并优化了检测大肠杆菌O157的二重荧光定量PCR方法,可检测的最低DNA浓度是10拷贝/μL;实验中O157菌株检测结果均为rfbE阳性,而非O157菌株检测结果均为阴性;重复性实验中,批间差异小于80%,批内差异小于70%。实验结果显示此二重荧光定量PCR方法可对分离的可疑大肠杆菌O157菌株进行快速鉴定,同时得知菌株是否携带stx2毒力基因,有利于菌株毒力强弱的判定。     

外来入侵植物粉花月见草的繁殖生物学特性

粉花月见草单花花期1d,整株花期50d左右;单花花粉量达8825个,花粉活力达53.2%,花粉萌发快,萌发率达44.6%,自花授粉;种子小,千粒重0.092g,种子发芽率达85%,萌发时间短,具有休眠机制。三年生株丛结果达1274个,单果种子高达206粒,单株丛种子量2.6×105。种子表面平滑,自然传播以重力为主和风力作用,传播距离在100cm范围内。结果表明:粉花月见草繁殖力、适应力强,易形成单一优势种群,种子小近圆形易形成种子库,人为活动使其远距离传播,具有较大的危害性。     

Distinct Expression Levels and Patterns of Stem Cell Marker,Aldehyde Dehydrogenase Isoform 1 (ALDH1), in Human Epithelial Cancers

Aldehyde dehydrogenase isoform 1 (ALDH1) has been proved useful for the identification of cancer stem cells. However, our knowledge of the expression and activity of ALDH1 in common epithelial cancers and their corresponding normal tissues is still largely absent. Therefore, we characterized ALDH1 expression in 24 types of normal tissues and a large collection of epithelial tumor specimens (six cancer types, n = 792) by immunohistochemical staining. Using the ALDEFUOR assay, ALDH1 activity was also examined in 16 primary tumor specimens and 43 established epithelial cancer cell lines. In addition, an ovarian cancer transgenic mouse model and 7 murine ovarian cancer cell lines were analyzed. We found that the expression levels and patterns of ALDH1 in epithelial cancers are remarkably distinct, and they correlate with their corresponding normal tissues. ALDH1 protein expression levels are positively correlated with ALDH1 enzymatic activity measured by ALDEFLUOR assay. Long-term in vitro culture doesn''t significantly affect ALDH1 activity in epithelial tumor cells. Consistent with research on other cancers, we found that high ALDH1 expression is significantly associated with poor clinical outcomes in serous ovarian cancer patients (n = 439, p = 0.0036). Finally, ALDH^br tumor cells exhibit cancer stem cell properties and are resistant to chemotherapy. As a novel cancer stem cell marker, ALDH1 can be used for tumors whose corresponding normal tissues express ALDH1 in relatively restricted or limited levels such as breast, lung, ovarian or colon cancer.     

Effects of neonatal isolation stress on hyperlocomotion of complexin II knockout mouse

The aim of this study was to investigate the influence of neonatal isolation stress on hyperlocomotion in complexin II knockout mouse (Cplx2(-/-)). The mice were randomly divided into 4 groups: Cplx2(-/-) with stress, Cplx2(+/+) with stress, Cplx2(-/-) without stress and Cplx2(+/+) without stress. Isolation stress was employed on the pups of stress groups from the 2nd day after the postnatal to the 21st day. The PCR was used to determine the gene type and the hyperlocomotion test was employed to detect the change of animal behavior after methamphetamine or saline injection (i.p.). The results showed that the animals of all groups increased their movement after injection of 0.2 mg/kg methamphetamine in different levels (P < 0.01), compared with those injected with saline. The Cplx2(-/-) mouse with stress revealed a significant increase in the distance of free movement after injection of 0.2 mg/kg methamphetamine compared with the knockout mouse without stress (P < 0.001). When Cplx2(-/-) mouse with stress was compared with wild type with stress, Cplx2(-/-) mouse with stress had more movement (P < 0.001), indicating that Cplx2 has effect on the hyperlocomotion as well. These results suggest an involvement of stress and Cplx2 in the movement behavior of mice.     

Protection of CHO cells by transfer of survivin driven by ovarian-specific promoter OSP-2

Chemotherapy is the major therapy for cancer in clinic. However, chemotherapeutic agents can harm the other tissues/organs besides cancer. Thus, there are great interests in protecting the innocents by the transfer of protective genes. There are two problems to be solved, one is the selection of protective genes and the other is the orientation of the exotic genes. Recent researches demonstrated that the principal mechanism of chemotherapeutics was through apoptosis. Hereby, introduction of anti-apoptosis genes might interrupt the processes of apoptosis to avoid side effect from chemotherapeutics. On the other hand, tissue-specific promoters, which control gene expression in a tissue-specific manner, might be an alternative tool to guarantee the location of target genes. In this research, we applied gene therapy to chemoprotection using anti-apoptosis gene survivin and ovarian-specific promoter OSP-2. The results showed that OSP-2 could specifically drive the expression of survivin in ovarian cells and survivin could protect cells via inhibiting apoptosis. This might put a light on the future of chemoprotective gene therapy.     

Influence of temperature and medium on viability of <Emphasis Type="Italic">Demodex folliculorum</Emphasis> and <Emphasis Type="Italic">Demodex brevis</Emphasis> (Acari: Demodicidae)

Demodex folliculorum and D. brevis are cosmopolitan, obligatory parasites. Epidemiological studies have established a clear association between these species and various facial diseases in humans. However, not much is known of the ecology of these mites, also because it has proven difficult to culture them. Here we report a laboratory study on the (combined) effects of temperature and culture medium on the viability of both species. Results showed significant differences in the survival time of D. folliculorum and D. brevis among different temperatures and media. Pair-wise comparisons revealed that both species survived longest in serum medium, at a temperature of 16–22°C. Therefore, we conclude that these conditions allow preservation and maintenance of both Demodex mites.     

Indications that live poultry markets are a major source of human H5N1 influenza virus infection in China

Human infections of H5N1 highly pathogenic avian influenza virus have continued to occur in China without corresponding outbreaks in poultry, and there is little conclusive evidence of the source of these infections. Seeking to identify the source of the human infections, we sequenced 31 H5N1 viruses isolated from humans in China (2005 to 2010). We found a number of viral genotypes, not all of which have similar known avian virus counterparts. Guided by patient questionnaire data, we also obtained environmental samples from live poultry markets and dwellings frequented by six individuals prior to disease onset (2008 and 2009). H5N1 viruses were isolated from 4 of the 6 live poultry markets sampled. In each case, the genetic sequences of the environmental and corresponding human isolates were highly similar, demonstrating a link between human infection and live poultry markets. Therefore, infection control measures in live poultry markets are likely to reduce human H5N1 infection in China.     

Methylenetetrahydrofolate reductase 677TT genotype might be associated with an increased lung cancer risk in Asians

Background

The association between methylenetetrahydrofolate reductase (MTHFR) 677C > T polymorphism and lung cancer risk has been studied in various populations with conflicting results. The aim of this study was to assess the association strength by a meta-analysis of published studies.

Methods

We searched PubMed and Chinese Biomedical (CBM) databases for relevant literatures published by July 18, 2012. Pooled odds ratio (OR) with 95% confidence interval (CI) was calculated to assess the strength of the association.

Results

A total of 20 studies comprising 11,653 cases and 12,032 controls were included in the final meta-analysis. Using the random effect model, we found that MTHFR 677TT variant genotype was associated with an increased lung cancer risk (OR = 1.26, 95% CI = 1.05–1.50, P = 0.011 for TT vs. CC; OR = 1.19, 95% CI = 1.03–1.37, P < 0.001 for TT vs. CC + CT; OR = 1.11, 95% CI = 1.02–1.22, P = 0.017 for T allele vs. C allele). In the further stratified analyses, the increased lung cancer risk was found in Asian subjects (OR = 1.31, 95% CI = 1.01–1.71, P = 0.045 for TT vs. CC; OR = 1.17, 95% CI = 1.00–1.38, P = 0.048 for TT vs. CC + CT). There were no evidences for obvious publication bias in the overall meta-analysis and Asian subjects.

Conclusions

MTHFR 677TT genotype might increase the susceptibility of lung cancer, especially in Asians.