... processed for RNA extraction RNA isolation, reversetranscriptionpolymerasechainreaction (RT- PCR) , andrealtimePCR RNA was extracted from frozen brains and cultured astrocytes with Trizol ... criteria of the Consortium to Establish a Registry for AD (CERAD) and NIA-Reagan guidelines Postmortem brain samples were obtained from male and female ND subjects and male and 11 female AD subjects ... hippocampus and cerebellum of AD and ND brains To demonstrate sPLA2-IIA mRNA expression in human brain, we measured levels of sPLA2-IIA mRNA by realtimePCR analysis of RNA prepared from hippocampus and...
... Methods Quantitative Reverse Transcriptase -Polymerase ChainReaction (RT- PCR) Real- time quantitative RT- PCR analysis of gene expression [13,14] was carried out on first-strand cDNA derived from ... seconds Real- time quantitative RT- PCR was assayed on an ABI Prism 7700 sequence detection system (Applied Biosystems, Foster City, CA) and the accumulation of PCR product was measured in realtime ... Teo IA and Shaunak S Polymerasechainreaction in situ: an appraisal of an emerging technique Histochem J 1995, 27:647-59 Heid CA, Stevens J, Livak KJ and Williams PM Realtime quantitative PCR...
... mẫu PolymeraseChainReaction Phản ứng chuỗi trùng hợp ReverseTranscriptionPolymeraseChainReaction Phản ứng chuỗi đồng phân hóa chép ngƣợc mRTPCR Multiplex ReverseTranscriptionPolymeraseChain ... đoán dựa nguyên lý PCR (RT- PCR realtime RT- PCR) Kỹ thuật PCR đƣợc phát triển để phát RNA virus PRRS mẫu bệnh phẩm Vì không cần phải phân lập virus môi trƣờng nuôi cấy tế bào nên PCR tốn thời gian ... phƣơng pháp nuôi cấy tế bào PCR đƣợc xem phƣơng pháp có độ nhậy độ đặc hiệu cao Phương pháp realtime RT- PCR Số hóa Trung tâm Học liệu http://www.lrc-tnu.edu.vn/ Realtime RT- PCR phƣơng pháp khuếch...
... NA của , virus cúm A/H5N1 thực hiê ̣n cùngmột phản ứng RT- PCR Phản ứng m utiplex RT- PCR: Được thực với sinh phẩm Qiagen OneStep RT- PCR theo hướng dẫn của nhà sả n xuấ t máy luân nhiê ... (H5N1) viruses and detection of anti-H5 antibody among household and social contacts”, J Infect Dis, 180, pp 1763-70 36 Lamb RA and Krug RM (2001), “Orthomyxoviridae: The Viruses and Their Replication”, ... mutiplex RT- PCR đươ ̣c thử nghiê ̣m với cả că ̣p mồ i để xác đinh các điề u kiê ̣n tố i ưu cho ̣ phản ứng Kế t quả các thử nghiê ̣m cho thấ y phản ứng m utiplex RT- PCR cho kế...
... virus isolationand real- time RT- PCR -/-: Negative results in both virus isolationand real- time RT- PCR 350 Dong Kun Yang et al has several advantages over conventional PCR First, realtime RT- PCR ... range of real- time RT- PCR were also evaluated When compared to the previous reports for detection of Flavivirus [7,16,25], real- time RT- PCR method Table Detection of JEV by real- time RT- PCRand virus ... TCID50 /reaction to 2.24 × 101.0 TCID50 /reaction Real- time RT- PCR assay could detect JE RNA ranging from 1.12 × 106.0 to 1.12 × 101.0 TCID50/ml Therefore, detection limits of RT- PCRand real- time RTPCR...
... observed by QC -RT- PCR it is unlikely that other methods to quantitate these mRNAs, e.g northern blotting or real- time PCR, would reveal a correlation either Conclusions QC -RT- PCRand related techniques ... Quantitative methods for reverse transcriptase polymerasechainreaction (RT- PCR) are rapidly surpassing all other methods of quantifying mRNA levels We therefore set out to develop an RT- PCR based method ... in a 25 µl PCR The remainder of the reactions were made up by µl PCR Buffer (Gibco BRL, Grand Island, NY), 0.25 µg of each primer, mM MgC12 in each CD4 reactionand mM MgC12 for CD8 reactions,...
... sinh học phân tử 20 1.3.3.1.Phương pháp RT – PCR (Reverse – transcriptionpolymerasechain reaction) 20 1.3.3.3 Các thành phần phản ứng PCR 23 CHƢƠNG 2: VẬT LIỆU VÀ PHƢƠNG PHÁP ... – PCR (Reverse – transcriptionpolymerasechain reaction) Phương pháp tạo RT- PCR gồm bước chính: - Tách chiết RNA từ bệnh phẩm biểu mô - Tạo cDNA phản ứng chép ngược - Thực phản ứng PCR với ... triphosphate dCTP: Deoxycytidine triphosphate UTRs: Untranslated regions RT- PCR: Reversetranscriptionpolymerasechainreaction kDa: kilo dalton H&E: Hematoxylin EoSin LỜI MỞ ĐẦU Ngày việc khai...
... with RT- PCRand nested PCR in the diagnosis of canine distemper Vet Res Commun 2005, 29:347-59 10 Li JZ, He HB, Xia XZ, et al.: Establishment and application of reversetranscriptionPolymerasechain ... chosen for the RT- nPCR Specificity of the multiplex RT- nPCR A fragment of 247 and 177 bp was amplified from CDV wild-type strain and the vaccine strain, respectively Two bands of 247 and 177 bp were ... which BioNote, Inc produced Multiplex reverse transcription- nested polymerasechainreaction Phylogenetic analysis and detection of CDV in field samples by RT- nPCR Total RNA was extracted from the...
... real- time RT- PCR of mRNA markers It shows the positive rate of mRNA markers from literature for detection of tumor cells by real- time RT- PCR Abbreviations RT- PCR: Reverse Transcriptase -Polymerase Chain ... carried out the real- time RT- PCR, participated in the clinical data collecting of the gastric carcinoma patients and drafted the manuscript LZH carried out the real- time RT- PCR ZZW participated in ... Analysis by Real- Time Quantitative PCR Quantitative PCR was performed using the Sequence Detector System, ABI PRISM 7500 (Applied Biosystems 7500 Fast Real- TimePCR System) PCR primers and the TaqMan...
... quantitative realtime RT- PCR Message expression in the synovium of knees and the relative differences in message levels between the control group and patients with OA or RA were determined by realtime RT- PCR ... Osteoarthritis Research Society [32] SD, standard deviation Table Sequences of the gene-specific oligonucleotide primers and probes for real- timereverse transcriptase polymerasechainreaction ... isoforms of HAS and hyaluronidase in knee synovium obtained from control donors and patients with OA or RA, by quantitative reverse transcriptase polymerasechainreaction (RT- PCR) , in order to...
... dilution was used for virus isolation, and a second aliquot was used for RNA extraction and duplex real- time RT- PCR analysis Table demonstrates that the duplex real- time RT- PCR assay was 10-fold more ... stored at -70°C, and the diluted working solutions were stored at -20°C [5] Sensitivity and specificity of the duplex real- time RT- PCR assay The sensitivity of the duplex real- time RT- PCR was conducted ... developed and validated a duplex realtime RT- PCR system for both WEEV and EEEV detection at the same time The primers and probes were selected from E3 gene for WEEV and NSP3 gene for EEEV, and the...
... pyrocarbonate-treated H2O and used as the template for the duplex real- time RT- PCR assay Duplex real- time RT- PCR amplification for HCV RNA detection The duplex real- time RT- PCR assay was performed ... which were detected by the duplex real- time RT- PCR assay (Table 4) Figure Linearity of the duplex real- time RT- PCR assay Linearity of the duplex real- time RT- PCR assay was determined using serial ... the duplex real- time RT- PCR for the 109 serum samples, which were identical to those of the CAP/CTM assay, should be correct The LOD of the duplex real- time RT- PCR assay was 38.99 IU/ml and the...
... Li Q, Porter BF, Barber RM, Claiborne MK, Levine JM, Levine GJ, Israel SK, Young BD, Kiupel M, et al.: Broadly reactive panparamyxovirus reversetranscriptionpolymerasechainreactionand sequence ... 21:844-849 21 Si W, Zhou S, Wang Z, Cui S: A multiplex reverse transcription- nested polymerasechainreaction for detection and differentiation of wildtype and vaccine strains of canine distemper virus ... Total RNA (1 μg) and random 8-mer primers (50 μM) were denatured at 65°C for and cooled down on ice To synthesise the first-strand cDNA, the RNA and primers were mixed in × reaction buffers,...
... virus However, single-step reverse transcriptase polymerasechainreaction (RT- PCR) detection and typing of dengue virus offers a sensitive, specific, reproducible and rapid alternative that ... minikit Reversetranscriptionandpolymerasechainreaction was performed using the One-Step Superscript III /RT/ Platinum Taq Mix (Invitrogen), 0.1 M dithiothreitol (DTT), 5' primer D1 and 3' primer ... being dengue positive and selected for confirmation and serotyping by reverse transcriptase polymerasechainreaction In 2003, 45 specimens were analysed of which 14 were males and 31 were females...
... 4°C Amplified Methylation Polymorphism PolymeraseChainReaction (AMP PCR) The PCRreaction consisted of DNA sample (genomic or digested template), Taq polymerase enzyme (AmpliTaq® Stoffel fragment, ... which were the HpaII recognition site (5’-CCGG-3’) and the other six bases were randomly designed (Table 1) The PCRreaction was started at 94°C for and each cycle was as follows: 94°C 30 sec, 57°C ... clear and reproducible marker bands were counted, and the comparison of bands between genomic and digested templates was done according to the appearance of the marker mentioned above Number and...
... thuật RT- PCR (Reverse Transcription - PolymeraseChain Reaction) Đây kỹ thuật sinh học phân tử, cho phép phát virus với độ xác cao, thời gian làm xét nghiệm khoảng 4-6 thực Realtime RT- PCR 12-24 ... Luria-Bertani media M: Matrix protein NA (N): Neuraminidase NP: Nucleoprotein NS: Non-structural protein PA: Polymerase A protein PB: Polymerase B protein RT- PCR: ReverseTranscription - PolymeraseChain ... khuôn mẫu để thực phản ứng PCR khuếch đại đoạn gen mong muốn Có phương pháp thực RT- PCR Đó phương pháp RT- PCR bước phương pháp RT- PCR hai bước Trong phương pháp RT- PCR hai bước, phản ứng phiên...
... endonuclease cleavage sites Polymerase chain reaction, cycle Primer Primer Template Cycle 1 Denature Anneal primers Synthesize new DNA with polymerasePolymerasechain reaction, cycle Cycle Denature ... produced on sucrose media: positive selection for recombinants PCR provides access to specific DNA segments • PolymeraseChainReaction • Requires knowledge of the DNA sequence in the region ... new DNA with polymerase PCR, cycle Cycle (focus on DNA segments bounded by primers) Denature Anneal primers Synthesize new DNA with polymerase duplex molecules of desired product PCR, cycle 4:...
... kỹ thuật PCR (Polymerase Chain Reaction) PCR phản ứng tổng hợp dây chuyền dùng để khuyếch đại trình tự DNA đích dựa bắt cặp đặc hiệu mồi với DNA mạch khn hoạt tính kéo dài cuả enzyme polymerase ... phản ứng, kích thước trình tự lai lực ion mơi trường 1.3.2.4 Phương pháp PCR (Polymerase Chain Reaction) Phương pháp PCR phương pháp in vitro để tổng hợp DNA dựa khn trình tự đích DNA ban đầu, ... ứng PCR - Hút 21l PCR mix bao gồm thành phần: PCR buffer (500mM KCl, 100mM Tris-HCl), MgCl2, dNTP, mồi, khn DNA - Cho thêm 3l DNA mẫu vừa xử lý nhiệt vào ống PCR - Bổ sung 1l Taq DNA polymerase...
... transcriptase PCR) gọi RNA – PCR hay RT – PCRRT – PCR nhạy phương pháp khác dùng cho phân tích ARN - RT – PCR cạnh tranh (Competitive RT – PCR) kỹ that thường dùng định lượng loại mARN chuyên biệt 21 - Real ... chuyên biệt 21 - Real – TimePCR đánh giá tích lũy sản phẩm định lượng qPCR (quantitative PCR – PCR định lượng) Ngoài số kỹ thuật PCR khác nữa… VIII RT – PCR Như biết, PCR kỹ thuật nhân bội ADN ... sản phẩm PCR Taq polymerase xúc tác tổng hợp, chúng kết hợp lại nhờ enzym ADN ligase Như sản phẩm PCR tách dòng vào vectơ T PCR hoàn thiện nhanh, nhiều biến dạng PCR đời : - RT – PCR (reverse...