PCR methods and applications最新文献

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Assessment of colinearity between large cloned DNA fragments and genomic DNA. 大克隆DNA片段与基因组DNA共线性的评估。
PCR methods and applications Pub Date : 1994-10-01 DOI: 10.1101/gr.4.2.129
P Verhasselt, G Volckaert
{"title":"Assessment of colinearity between large cloned DNA fragments and genomic DNA.","authors":"P Verhasselt, G Volckaert","doi":"10.1101/gr.4.2.129","DOIUrl":"https://doi.org/10.1101/gr.4.2.129","url":null,"abstract":"","PeriodicalId":77315,"journal":{"name":"PCR methods and applications","volume":"4 2","pages":"129-32"},"PeriodicalIF":0.0,"publicationDate":"1994-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"18585845","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 3
Detection of p53 gene mutation in cancer tissues by nonradioactive direct sequencing. 非放射性直接测序法检测癌组织中p53基因突变。
PCR methods and applications Pub Date : 1994-10-01 DOI: 10.1101/gr.4.2.76
D Bautista, P Chaubert, J Bertoncini, J Benhattar
{"title":"Detection of p53 gene mutation in cancer tissues by nonradioactive direct sequencing.","authors":"D Bautista,&nbsp;P Chaubert,&nbsp;J Bertoncini,&nbsp;J Benhattar","doi":"10.1101/gr.4.2.76","DOIUrl":"https://doi.org/10.1101/gr.4.2.76","url":null,"abstract":"<p><p>A simple procedure for direct sequencing of double-stranded PCR products by the dideoxy-termination method has been developed using biotinylated sequencing primers. Sequences of the p53 gene have been obtained from DNA extracted from frozen and formalin-fixed paraffin-embedded cancer tissues. Detection of sequencing ladders was done with chemiluminescent or colorimetric techniques. Both are highly sensitive, but colorimetric detection is less prone to diffusion artifacts, which are common in G-rich regions. Use of this nonradioactive PCR-sequencing protocol allowed rapid and simple determination of p53 gene alterations in human tumors.</p>","PeriodicalId":77315,"journal":{"name":"PCR methods and applications","volume":"4 2","pages":"76-9"},"PeriodicalIF":0.0,"publicationDate":"1994-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"18587059","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 7
Analysis of nonspecific DNA synthesis during in situ PCR and solution-phase PCR. 原位PCR和液相PCR非特异性DNA合成分析。
PCR methods and applications Pub Date : 1994-10-01 DOI: 10.1101/gr.4.2.89
G J Nuovo, P MacConnell, F Gallery
{"title":"Analysis of nonspecific DNA synthesis during in situ PCR and solution-phase PCR.","authors":"G J Nuovo,&nbsp;P MacConnell,&nbsp;F Gallery","doi":"10.1101/gr.4.2.89","DOIUrl":"https://doi.org/10.1101/gr.4.2.89","url":null,"abstract":"<p><p>In this study we examine the factors that lead to nonspecific DNA synthesis during in situ PCR and solution-phase PCR. It was shown that primer-independent DNA synthesis can produce an intense signal during in situ PCR. This primer-independent pathway was apparently the result of the repair of DNA gaps induced by the heat treatment of the paraffin embedded tissue sections. This non-specific signal could be eliminated by blocking gap repair with dideoxy-TTP, avoiding heat treatment, or DNase pretreatment. The primer-independent signal was also influenced by the length and mode of fixation and the sample tissue itself. Elimination of the primer-independent signal and the use of viral primers in tissues that did not contain the virus showed that nonspecific DNA synthesis could be eliminated by the hot start modification. Primer oligomerization did not produce a signal during in situ PCR, even when it occurred robustly in the amplifying solution. Generation of the primer-independent signal in solution-phase PCR with purified DNA required a cross-linking fixative, heating, the addition of bovine serum albumin, and intact protein-DNA cross-links.</p>","PeriodicalId":77315,"journal":{"name":"PCR methods and applications","volume":"4 2","pages":"89-96"},"PeriodicalIF":0.0,"publicationDate":"1994-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"18587062","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 18
PCR analysis of the H ferritin multigene family reveals the existence of two classes of processed pseudogenes. 对H铁蛋白多基因家族的PCR分析显示存在两类加工假基因。
PCR methods and applications Pub Date : 1994-10-01 DOI: 10.1101/gr.4.2.85
B Quaresima, M T Tiano, A Porcellini, P D'Agostino, M C Faniello, M A Bevilacqua, F Cimino, F Costanzo
{"title":"PCR analysis of the H ferritin multigene family reveals the existence of two classes of processed pseudogenes.","authors":"B Quaresima,&nbsp;M T Tiano,&nbsp;A Porcellini,&nbsp;P D'Agostino,&nbsp;M C Faniello,&nbsp;M A Bevilacqua,&nbsp;F Cimino,&nbsp;F Costanzo","doi":"10.1101/gr.4.2.85","DOIUrl":"https://doi.org/10.1101/gr.4.2.85","url":null,"abstract":"<p><p>The human gene coding for the apoferritin H subunit belongs to a complex multigene family constituted by the expressed gene and by an undefined number of pseudogenes. We have used a strategy based on PCR to amplify specifically the H pseudogenes from a sample of human genomic DNA. With this approach, three new H pseudogenes have been cloned and characterized by DNA sequence analysis. In addition, we have identified a new type of pseudogene, the size of which (700 bp) is caused by multiple detection events in the putative coding region.</p>","PeriodicalId":77315,"journal":{"name":"PCR methods and applications","volume":"4 2","pages":"85-8"},"PeriodicalIF":0.0,"publicationDate":"1994-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"18587061","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 8
Rapid isolation of cDNA clones by aliquot testing via PCR amplification. PCR扩增等分检测快速分离cDNA克隆。
PCR methods and applications Pub Date : 1994-10-01 DOI: 10.1101/gr.4.2.126
A Di Bacco, L Susani, A Villa, D Strina, A Frattini, P Vezzoni, I Zucchi
{"title":"Rapid isolation of cDNA clones by aliquot testing via PCR amplification.","authors":"A Di Bacco,&nbsp;L Susani,&nbsp;A Villa,&nbsp;D Strina,&nbsp;A Frattini,&nbsp;P Vezzoni,&nbsp;I Zucchi","doi":"10.1101/gr.4.2.126","DOIUrl":"https://doi.org/10.1101/gr.4.2.126","url":null,"abstract":"","PeriodicalId":77315,"journal":{"name":"PCR methods and applications","volume":"4 2","pages":"126-8"},"PeriodicalIF":0.0,"publicationDate":"1994-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"18585844","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 2
Expression-PCR (E-PCR): overview and applications. 表达pcr (E-PCR)技术综述及应用
PCR methods and applications Pub Date : 1994-10-01 DOI: 10.1101/gr.4.2.s92
D E Lanar, K C Kain
{"title":"Expression-PCR (E-PCR): overview and applications.","authors":"D E Lanar,&nbsp;K C Kain","doi":"10.1101/gr.4.2.s92","DOIUrl":"https://doi.org/10.1101/gr.4.2.s92","url":null,"abstract":"","PeriodicalId":77315,"journal":{"name":"PCR methods and applications","volume":"4 2","pages":"S92-6"},"PeriodicalIF":0.0,"publicationDate":"1994-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"18585839","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 17
Detection and differential display of expressed genes by DDRT-PCR. 表达基因的DDRT-PCR检测及差异显示。
PCR methods and applications Pub Date : 1994-10-01 DOI: 10.1101/gr.4.2.s97
D Bauer, P Warthoe, M Rohde, M Strauss
{"title":"Detection and differential display of expressed genes by DDRT-PCR.","authors":"D Bauer,&nbsp;P Warthoe,&nbsp;M Rohde,&nbsp;M Strauss","doi":"10.1101/gr.4.2.s97","DOIUrl":"https://doi.org/10.1101/gr.4.2.s97","url":null,"abstract":"","PeriodicalId":77315,"journal":{"name":"PCR methods and applications","volume":"4 2","pages":"S97-108"},"PeriodicalIF":0.0,"publicationDate":"1994-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"18585840","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 44
Effect of internal direct and inverted Alu repeat sequences on PCR. 内部直接和反向Alu重复序列对PCR的影响。
PCR methods and applications Pub Date : 1994-10-01 DOI: 10.1101/gr.4.2.109
W Ji, X Y Zhang, G S Warshamana, G Z Qu, M Ehrlich
{"title":"Effect of internal direct and inverted Alu repeat sequences on PCR.","authors":"W Ji,&nbsp;X Y Zhang,&nbsp;G S Warshamana,&nbsp;G Z Qu,&nbsp;M Ehrlich","doi":"10.1101/gr.4.2.109","DOIUrl":"https://doi.org/10.1101/gr.4.2.109","url":null,"abstract":"<p><p>We have studied the effect of repeated DNA sequence, especially Alu repeats, on PCR. Alu repeats are sequences that are approximately 300 bp long and interspersed at a very high copy number throughout the human genome. We amplified part of the human low-density lipoprotein receptor gene containing two Alu repeat sequences in the same orientation, approximately 7.8 kb apart, with unique sequence primers outside these repeats. The major PCR product was a DNA fragment with an in vitro deletion between the Alu repeats. The formation of this product depended on the template concentration and the type of polymerase used. Such a product arose apparently as a result of a \"jumping reaction\" involving a primer whose extension was terminated prematurely within one Alu repeated followed by annealing of such an incompletely extended primer to the other, distant Alu repeat. No such jumping products were seen when a 0.8-kb region containing two nearby inverted Alu repeats within the human alpha-galactosidase A gene was subject to PCR with unique sequence primers annealing just outside these repeats.</p>","PeriodicalId":77315,"journal":{"name":"PCR methods and applications","volume":"4 2","pages":"109-16"},"PeriodicalIF":0.0,"publicationDate":"1994-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"18585841","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 11
Use of vectorette and subvectorette PCR to isolate transgene flanking DNA. 利用载体和亚载体PCR分离转基因侧翼DNA。
PCR methods and applications Pub Date : 1994-10-01 DOI: 10.1101/gr.4.2.71
M J Allen, A Collick, A J Jeffreys
{"title":"Use of vectorette and subvectorette PCR to isolate transgene flanking DNA.","authors":"M J Allen,&nbsp;A Collick,&nbsp;A J Jeffreys","doi":"10.1101/gr.4.2.71","DOIUrl":"https://doi.org/10.1101/gr.4.2.71","url":null,"abstract":"<p><p>Vectorette PCR permits the specific amplification of DNA segments flanking a known DNA sequence. It enables the application of the PCR where sequence information is only available for one primer site. We now show that vectorette PCR can be used for the systematic mapping and retrieval of transgene flanking DNA. We also show that the sequence of large vectorette PCR fragments can be obtained without cloning, by the production of subvectorette fragments.</p>","PeriodicalId":77315,"journal":{"name":"PCR methods and applications","volume":"4 2","pages":"71-5"},"PeriodicalIF":0.0,"publicationDate":"1994-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"18587058","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 27
Computing genetic similarity coefficients from RAPD data: correcting for the effects of PCR artifacts caused by variation in experimental conditions. 从RAPD数据计算遗传相似系数:校正由实验条件变化引起的PCR伪影的影响。
PCR methods and applications Pub Date : 1994-08-01 DOI: 10.1101/gr.4.1.38
W F Lamboy
{"title":"Computing genetic similarity coefficients from RAPD data: correcting for the effects of PCR artifacts caused by variation in experimental conditions.","authors":"W F Lamboy","doi":"10.1101/gr.4.1.38","DOIUrl":"https://doi.org/10.1101/gr.4.1.38","url":null,"abstract":"<p><p>The production of informative random amplified polymorphic DNA (RAPD) markers using PCR and a single primer is often accompanied by the generation of artifactual (noninformative) bands as well. When RAPD data are used to compute genetic similarity coefficients, these artifacts (false positives, false negatives, or both) can cause large biases in the numerical values of the coefficients. As a result, some workers have been reluctant to use RAPD markers in the estimation of genetic similarities. Artifactual bands are of two types: those caused by variation in experimental conditions, and those caused by characteristics of the DNA to be amplified. A procedure is described that allows for correction of the bias caused by the first type of artifact, providing that replicate DNA samples have been extracted, amplified, and scored. The resulting data are used to obtain an estimate of the proportion of false-positive and false-negatives bands. These values are then used to correct the bias in the computed similarity coefficients. Two examples are given, one in which bias correction is critical to the results, and one in which it is less important. The maximum percent bias, computed from the estimated proportions of false positives and false negatives in the RAPD data set, is proposed as a criterion for determining whether bias correction of the similarity coefficients is required or not. Although all reasonable efforts should be made to optimize PCR protocols to eliminate artifactual bands, when this is not possible, the methods described allow RAPD markers to compute genetic similarities reliably and accurately, even when artifactual bands resulting from variation in experimental conditions are present.</p>","PeriodicalId":77315,"journal":{"name":"PCR methods and applications","volume":"4 1","pages":"38-43"},"PeriodicalIF":0.0,"publicationDate":"1994-08-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"19977478","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 36
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