Decontamination of DNA in Taq DNA polymerase reagents using nylon membranes for monitoring of GMOs

Zhao Shi Rong,Qu Bo,Kim Jae Kwang,Lee Bumkyu 한국식물생명공학회 2021 Plant biotechnology reports Vol.15 No.6

Polymerase chain reaction (PCR) is one of the most commonly used methods in diagnostic and molecular biology. However, DNA contamination can lead to incorrect results. This poses a major problem, especially in the monitoring of genetically modified organisms (GMOs). We investigated DNA contamination in 16 commercial Taq DNA polymerase reagents. Many of the Taq DNA polymerase reagents were contaminated with the 16S rRNA gene and an ampicillin resistance gene (bla). We attempted to decontaminate the Taq DNA polymerase reagents using UV, γ-ray, and electron-ray irradiation, as well as nylon membrane and FTA card treatment. UV irradiation reduced the activity of Taq DNA polymerase. Various wavelengths of γ-ray and electron-ray irradiation had no decontamination effect. However, 24-h treatment with three nylon membrane disks removed DNA contamination. FTA card treatment, which has membrane-like functions, also removed decontamina- tion but decreased enzyme activity. We further confirmed that the nylon membrane-treated Taq DNA polymerase reagent could be used in the detection of the bla and NPTII genes in genetically modified E. coli. These results are useful for future monitoring of GMOs.

Metastasis associated genomic aberrations in stage II rectal cancer

Hong Zhao,Zhi-Zhou Shi,Rui Jiang,Dong-Bing Zhao,Hai-Tao Zhou,Jian-Wei Liang,Xin-Yu Bi,Jian-Jun Zhao,Zhi-Yu Li,Jian-Guo Zhou,Zhen Huang,Ye-Fan Zhang,Jian Wang,Xin Xu,Yan Cai,Ming-Rong Wang,Yu Zhang 한국유전학회 2016 Genes & Genomics Vol.38 No.11

Genomic aberrations of rectal carcinoma, especially DNA copy number changes associated with metastasis were largely unclear. We aim to identify the metastasis associated biomarkers in stage II rectal cancer. Formalin-fixed, paraffin-embedded primary tumor tissues of stage II rectal carcinoma were analyzed by array-based comparative genomic hybridization, and genomic aberrations were identified by Genomic Workbench and SAM software. Copy number changes and mRNA expressions were validated by Real-time PCR in an independent rectal cancer samples. The results showed that the most frequent gains in stage II rectal cancer were at 1q21.2-q23.1, 3p21.31, 11q12.2-q23.3, 12q24.11-q24.31, 12q13.11-q14.1 and losses in 18q11.2-q23, 17q21.33-q22, 13q31.1-q31.3, 21q21.1-q21.3, 8p23.3-p23.1 and 4q22.1-q23. Twenty-two amplifications and five homozygous deletions were also identified. We further found that S100A1 (1q21.3-q23.1), MCM7 (7q22.1) and JUND (19p13.11) were amplified and overexpressed in stage II rectal cancer. Interestingly, the genomic aberrations affected 14 signaling pathways including VEGF signaling pathway and fatty acid metabolism. Most importantly, loss of 13q31.1-q34 and gain of 1q44 were associated with distant metastasis. Our results indicated that these metastasis associated genomic changes may be useful to reveal the pathogenesis of rectal cancer metastasis and identify candidate biomarkers.

Behaviour of FRP composite columns: Review and analysis of the section forms

Rong, Chong,Shi, Qingxuan,Zhao, Hongchao Techno-Press 2020 Advances in concrete construction Vol.9 No.2

As confining materials for concrete, steel and fibre-reinforced polymer (FRP) composites have important applications in both the seismic retrofit of existing reinforced concrete columns and in the new construction of composite structures. We present a comprehensive review of the axial stress-strain behaviour of the FRP-confined concrete column. Next, the mechanical performance of the hybrid FRP-confined concrete-steel composite columns are comprehensively reviewed. Furthermore, the results of FRP-confined concrete column experiments and FRP-confined circular concrete-filled steel tube experiments are presented to study the interaction relationship between various material sections. Finally, the combinations of material sections are discussed. Based on these observations, recommendations regarding future research directions for composite columns are also outlined.

8q24 rs4242382 Polymorphism is a Risk Factor for Prostate Cancer among Multi-Ethnic Populations: Evidence from Clinical Detection in China and a Meta-analysis

Zhao, Cheng-Xiao,Liu, Ming,Xu, Yong,Yang, Kuo,Wei, Dong,Shi, Xiao-Hong,Yang, Fan,Zhang, Yao-Guang,Wang, Xin,Liang, Si-Ying,Zhao, Fan,Zhang, Yu-Rong,Wang, Na-Na,Chen, Xin,Sun, Liang,Zhu, Xiao-Quan,Yuan Asian Pacific Journal of Cancer Prevention 2014 Asian Pacific journal of cancer prevention Vol.15 No.19

Background: Evidence supporting an association between the 8q24 rs4242382-A polymorphism and prostate cancer (PCa) risk has been reported in North American and Europe populations, though data from Asian populations remain limited. We therefore investigated this association by clinical detection in China, and meta-analysis in Asian, Caucasian and African-American populations. Materials and Methods: Blood samples and clinical information were collected from ethnically Chinese men from Northern China with histologically-confirmed PCa (n=335) and from age-matched normal controls (n=347). The 8q24 (rs4242382) gene polymorphism was genotyped by polymerase chain reaction-high-resolution melting analysis. We initially analyzed the associations between the risk allele and PCa and clinical covariates. A meta-analysis was then performed using genotyping data from a total of 1,793 PCa cases and 1,864 controls from our study and previously published studies in American and European populations, to determine the association between PCa and risk genotype. Results: The incidence of the risk allele was higher in PCa cases than controls (0.222 vs 0.140, $P=7.3{\times}10^{-5}$), suggesting that the 8q24 rs4242382-A polymorphism was associated with PCa risk in Chinese men. The genotypes in subjects were in accordance with a dominant genetic model (ORadj=2.03, 95%CI: 1.42-2.91, $Padj=1.1{\times}10^{-4}$). Presence of the risk allele rs4242382-A at 8q24 was also associated with clinical covariates including age at diagnosis ${\geq}65$ years, prostate specific antigen >10 ng/ml, Gleason score <8, tumor stage and aggressive PCa, compared with the non-risk genotype ($P=4.6{\times}10^{-5}-3.0{\times}10^{-2}$). Meta-analysis confirmed the association between 8q24 rs4242382-A polymorphism and PCa risk (OR=1.62, 95%CI: 1.39-1.88, $P=1.0{\times}10^{-5}$) across Asian, Caucasian and African American populations. Conclusions: The replicated data suggest that the 8q24 rs4242382-A variation might be associated with increased PCa susceptibility in Asian, Caucasian and African American populations. These results imply that this polymorphism may be a useful risk biomarker for PCa in multi-ethnic populations.

MicroRNA-27a Inhibits Cell Migration and Invasion of Fibroblast-Like Synoviocytes by Targeting Follistatin-Like Protein 1 in Rheumatoid Arthritis

Shi, Dong-liang,Shi, Gui-rong,Xie, Jing,Du, Xu-zhao,Yang, Hao Korean Society for Molecular and Cellular Biology 2016 Molecules and cells Vol.39 No.8

Fibroblast-like synoviocytes (FLS) with aberrant expression of microRNA (miRNA) are critical pathogenic regulators in rheumatoid arthritis (RA). Previous studies have found that overexpression or silencing of miRNA can contribute to the development of miRNA-based therapeutics in arthritis models. In this study, we explored the effects of miR-27a on cell migration and invasion in cultured FLS from RA patients. We found that miR-27a was markedly downregulated in the serum, synovial tissue, and FLS of RA patients. Meanwhile, the expression of follistatin-like protein 1 (FSTL1) was upregulated, which suggests that FSTL1 plays a key role in RA development. The results of a Transwell assay showed that miR-27a inhibited FLS migration and invasion. However, miR-27a inhibition promoted the migration and invasion of FLS. In addition, the down-regulated expression of matrix metalloproteinases (MMP2, MMP9, and MMP13) and Rho family proteins (Rac1, Cdc42, and RhoA) was detected after treatment with miR-27a in RA-FLS by quantitative reverse transcription-PCR and western blot analysis. Then, a luciferase reporter assay validated that miR-27a targeted the 3-untranslated region (3'-UTR) of FSTL1. Moreover, miR-27a caused a significant decrease of FSTL1. In addition, the expression of TLR4 and $NF{\kappa}B$ was inhibited by miR-27a but increased by FSTL1 overexpression. In conclusion, we found that miR-27a inhibited cell migration and invasion of RA-FLS by targeting FSTL1 and restraining the $TLR4/NF{\kappa}B$ pathway.

Microarrays for the Detection of HBV and HDV

( Zhao Hui Sun ),( Wen Ling Zheng ),( Bao Zhang ),( Rong Shi ),( Wen Li Ma ) 생화학분자생물학회 2004 BMB Reports Vol.37 No.5

The increasing pace of development in molecular biology during the last decade has had a direct effect on mass testing and diagnostic applications, including blood screening. We report the model Microarray that has been developed for Hepatitis B virus (HBV) and Hepatitis D virus (HDV) detection. The specific primer pairs of PCR were designed using the Primer Premier 5.00 program according to the conserved regions of HBV and HDV. PCR fragments were purified and cloned into pMD18-T vectors. The recombinant plasmids were extracted from positive clones and the target gene fragments were sequenced. The DNA microarray was prepared by robotically spotting PCR products onto the surface of glass slides. Sequences were aligned, and the results obtained showed that the products of PCR amplification were the required specific gene fragments of HBV, and HDV. Samples were labeled by Restriction Display PCR (RD-PCR). Gene chip hybridizing signals showed that the specificity and sensitivity required for HBV and HDV detection were satisfied. Using PCR amplified products to construct gene chips for the simultaneous clinical diagnosis of HBV and HDV resulted in a quick, simple, and effective method. We conclude that the DNA microarray assay system might be useful as a diagnostic technique in the clinical laboratory. Further applications of RD-PCR for the sample labeling could speed up microarray multi-virus detection.

Interoperative Radiotherapy of Seventy-two Cases of Early Breast Cancer Patients During Breast-conserving Surgery

Zhou, Shi-Fu,Shi, Wei-Feng,Meng, Dong,Sun, Chun-Lei,Jin, Jian-Rong,Zhao, Yu-Tian Asian Pacific Journal of Cancer Prevention 2012 Asian Pacific journal of cancer prevention Vol.13 No.4

Objective: To evaluate interoperative radiotherapy after breast conservative surgery in early breast cancer patients in terms of postoperative complications, cosmetic outcome and recurrence events. Methods: From June 2007 to Dec 2011, 143 early breast cancer patients received breast conservative surgery. Seventy-two (study group) received interoperative radiotherapy, compared with 71 patients (control group) given routine radiotherapy. Postoperative complications were evaluated 1 month after surgery; cosmetic outcome was evaluated 1 year postoperatively; recurrence and death events were followed up. Results: The average wound healing time was 13~22 d in the study group and 9~14 d in the control group. In the study group, 2 patients developed lyponecrosis, 16 patients showed wound edema while no such side effects were found in the control group. No infection or hematomas were found in either group. In the study group (59 cases), overall cosmetic outcome in 53 patients was graded as excellent or good, and in 6 as fair or poor. Meanwhile in the control group (56 cases), 42 patients were graded as excellent or good, and 14 as fair or poor (P=0.032). After a follow-up from 3 to 54 months (median: 32 months), two patients (2.78%) in study group developed local relapses, one of them (1.39%) died, 2 patients (2.78%) developed bone metastases. In control group, one patient (1.41%) developed local relapse, 2 patients (2.82%) developed bone metastases, and no one died. Conclusion: Intraoperative radiotherapy is safe and reliable with good cosmetic outcome.

Value of PAX1 Methylation Analysis by MS-HRM in the Triage of Atypical Squamous Cells of Undetermined Significance

Li, Shi-Rong,Wang, Zhen-Ming,Wang, Yu-Hui,Wang, Xi-Bo,Zhao, Jian-Qiang,Xue, Hai-Bin,Jiang, Fu-Guo Asian Pacific Journal of Cancer Prevention 2015 Asian Pacific journal of cancer prevention Vol.16 No.14

Background: Detection of cervical high grade lesions in patients with atypical squamous cells of undetermined significance (ASCUS) is still a challenge. Our study tested the efficacy of the paired boxed gene 1 (PAX1) methylation analysis by methylation-sensitive high-resolution melting (MS-HRM) in the detection of high grade lesions in ASCUS and compared performance with the hybrid capture 2 (HC2) human papillomavirus (HPV) test. Materials and Methods: A total of 463 consecutive ASCUS women from primary screening were selected. Their cervical scrapings were collected and assessed by PAX1 methylation analysis (MS-HRM) and high-risk HPV-DNA test (HC2). All patients with ASCUS were admitted to colposcopy and cervical biopsies. The Chisquare test was used to test the differences of PAX1 methylation or HPV infection between groups. Results: The specificity, sensitivity, and accuracy for detecting CIN2 + lesions were: 95.6%, 82.4%, and 94.6%, respectively, for the PAX1 MS-HRM test; and 59.7%, 64.7%, and 60.0% for the HC2 HPV test. Conclusions: The PAX1 methylation analysis by MS-HRM demonstrated a better performance than the high-risk HPV-DNA test for the detection of high grade lesions (CIN2 +) in ASCUS cases. This approach could screen out the majority of low grade cases of ASCUS, and thus reduce the referral rate to colposcopy.

Crashworthy and Performance-cost Characteristics of Aluminum-CFRP Hybrid Tubes under Quasi-static Axial Loading

Peilong Shi,Qiang Yu,Rong Huang,Xuan Zhao,Guohua Zhu 한국섬유공학회 2019 Fibers and polymers Vol.20 No.2

Metal/composite hybrid thin-walled structures combine the low cost of metallic materials and the high strength-toweight ratio of composites and thus have the potential to be utilized as cost-effective energy absorbers for vehicle applications. This study aimed to examine the crushing behaviors and performance-to-cost ratio of aluminum/carbon fiberreinforced plastic (CFRP) hybrid tubes under quasi-static axial loading. First, a single aluminum tube, a single CFRP tube and an aluminum/CFRP hybrid tube were tested to validate numerical models. The experimental results showed that the total energy absorption (EA) of the aluminum/CFRP hybrid tube was 32.46 % higher than the sum of that of the individual parts, and the special energy absorption (SEA) of the hybrid tube was improved by 105.26 % compared with that of the single aluminum tube. Then, the effects of the orientation angles ([±15 o]n, [±45 o]n, [±75 o]n, [±90 o]n, [90 o/0 o]n, n=2, 4, 6) and thicknesses of the CFRP tube wall (4-ply, 8-ply, 12-ply) on the crashworthiness of the hybrid tube were studied through validated numerical models. The numerical results showed that as the orientation angle increased, SEA, EA and the mean crushing force (Fmean) decreased first and then increased; in addition, the hybrid tubes with orientation angles of [±45 o]n and [90 o/0 o]n (n=2, 4, 6) consistently exhibited the worst and best crashworthiness, respectively. Furthermore, the SEA, EA, and Fmean of the hybrid tube increased with increasing thickness of the CFRP tube wall. Finally, the performance-to-cost ratio (SEA/cost) of the hybrid tube was analyzed, and the results show that aluminum/CFRP hybrid tubes with a smaller wall thickness of the CFRP tube exhibits superior potential in terms of both cost and performance for automotive applications.

Expression Profile and Potential Roles of EVA1A in Normal and Neoplastic Pancreatic Tissues

Tao, Ming,Shi, Xue-Ying,Yuan, Chun-Hui,Hu, Jia,Ma, Zhao-Lai,Jiang, Bin,Xiu, Dian-Rong,Chen, Ying-Yu Asian Pacific Journal of Cancer Prevention 2015 Asian Pacific journal of cancer prevention Vol.16 No.1

Background: EVA1A (eva-1 homolog A) is a novel gene that regulates programmed cell death through autophagy and apoptosis. Our objective was to investigate the expression profiles and potential role of EVA1A in normal and neoplastic human pancreatic tissues. Materials and Methods: The expression pattern of EVA1A in normal pancreatic tissue was examined by indirect immunofluorescence and confocal microscopy. Protein levels in paraffin-embedded specimens from normal and diseased pancreatic and matched non-tumor tissues were evaluated by immunohistochemistry. Results: EVA1A colocalized with glucagon but not with insulin, demonstrating production in islet alpha cells. Itwas strongly expressed in chronic pancreatitis, moderately or weakly expressed in the plasma membrane and cytoplasm in pancreatic acinar cell carcinoma, and absent in normal pancreatic acinar cells. Although the tissue architecture was deformed, EVA1A was absent in the alpha cells of pancreatic ductal adenocarcinomas, intraductal papillary mucinous neoplasms, mucinous cystadenomas, solid papillary tumors and pancreatic neuroendocrine tumors. Conclusions: EVA1A protein is specifically expressed in islet alpha cells, suggesting it may play an important role in regulating alpha-cell function. The ectopic expression of EVA1A in pancreatic neoplasms may contribute to their pathogenesis and warrants further investigation.