H J Lenz, C Hill, K D Danenberg, L L Leichman, D G Priest, P V Danenberg
{"title":"Rapid quantitative PCR for determination of relative gene expressions in tissue specimens.","authors":"H J Lenz, C Hill, K D Danenberg, L L Leichman, D G Priest, P V Danenberg","doi":"10.1101/gr.4.5.305","DOIUrl":"https://doi.org/10.1101/gr.4.5.305","url":null,"abstract":"","PeriodicalId":77315,"journal":{"name":"PCR methods and applications","volume":"4 5","pages":"305-8"},"PeriodicalIF":0.0,"publicationDate":"1995-04-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"18586922","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}
{"title":"Panhandle PCR.","authors":"D H Jones","doi":"10.1101/gr.4.5.s195","DOIUrl":"https://doi.org/10.1101/gr.4.5.s195","url":null,"abstract":"","PeriodicalId":77315,"journal":{"name":"PCR methods and applications","volume":"4 5","pages":"S195-201"},"PeriodicalIF":0.0,"publicationDate":"1995-04-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"18586342","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}
A Bobba, R Lippolis, S Giannattasio, C Camaschella, E Marra
{"title":"An efficient method for PCR analysis of mitochondrial DNA from paraffin-embedded archival heart tissue.","authors":"A Bobba, R Lippolis, S Giannattasio, C Camaschella, E Marra","doi":"10.1101/gr.4.5.309","DOIUrl":"https://doi.org/10.1101/gr.4.5.309","url":null,"abstract":"","PeriodicalId":77315,"journal":{"name":"PCR methods and applications","volume":"4 5","pages":"309-10"},"PeriodicalIF":0.0,"publicationDate":"1995-04-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"18586923","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}
We have designed a novel strategy for the isolation of the rearranged genomic fragments encoding the L-VH-D-JH and L-V kappa/lambda-J kappa/lambda regions of mouse immunoglobulin genes. This strategy is based on the PCR amplification of genomic DNA from mouse hybridomas using multiple specific primers chosen in the 5'-untranslated region and in the intron downstream of the rearranged JH/J kappa/lambda sequences. Variable regions with intact coding sequences, including full-length leader peptides (L) can be obtained without previous DNA sequencing. Our strategy is based on a genomic template that produces fragments that do not need to be adapted for recombinant antibody expression, thus facilitating the generation of chimeric and isotype-switched immunoglobulins.
{"title":"Specific amplification by PCR of rearranged genomic variable regions of immunoglobulin genes from mouse hybridoma cells.","authors":"J Berdoz, T P Monath, J P Kraehenbuhl","doi":"10.1101/gr.4.5.256","DOIUrl":"https://doi.org/10.1101/gr.4.5.256","url":null,"abstract":"<p><p>We have designed a novel strategy for the isolation of the rearranged genomic fragments encoding the L-VH-D-JH and L-V kappa/lambda-J kappa/lambda regions of mouse immunoglobulin genes. This strategy is based on the PCR amplification of genomic DNA from mouse hybridomas using multiple specific primers chosen in the 5'-untranslated region and in the intron downstream of the rearranged JH/J kappa/lambda sequences. Variable regions with intact coding sequences, including full-length leader peptides (L) can be obtained without previous DNA sequencing. Our strategy is based on a genomic template that produces fragments that do not need to be adapted for recombinant antibody expression, thus facilitating the generation of chimeric and isotype-switched immunoglobulins.</p>","PeriodicalId":77315,"journal":{"name":"PCR methods and applications","volume":"4 5","pages":"256-64"},"PeriodicalIF":0.0,"publicationDate":"1995-04-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1101/gr.4.5.256","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"18586913","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}
S Cheng, Y Chen, J A Monforte, R Higuchi, B Van Houten
{"title":"Template integrity is essential for PCR amplification of 20- to 30-kb sequences from genomic DNA.","authors":"S Cheng, Y Chen, J A Monforte, R Higuchi, B Van Houten","doi":"10.1101/gr.4.5.294","DOIUrl":"https://doi.org/10.1101/gr.4.5.294","url":null,"abstract":"","PeriodicalId":77315,"journal":{"name":"PCR methods and applications","volume":"4 5","pages":"294-8"},"PeriodicalIF":0.0,"publicationDate":"1995-04-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1101/gr.4.5.294","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"18586919","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}
{"title":"Optimization and troubleshooting in PCR.","authors":"K H Roux","doi":"10.1101/gr.4.5.s185","DOIUrl":"https://doi.org/10.1101/gr.4.5.s185","url":null,"abstract":"","PeriodicalId":77315,"journal":{"name":"PCR methods and applications","volume":"4 5","pages":"S185-94"},"PeriodicalIF":0.0,"publicationDate":"1995-04-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"18586341","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}
{"title":"Efficient gene synthesis by Klenow assembly/extension-Pfu polymerase amplification (KAPPA) of overlapping oligonucleotides.","authors":"E W Holowachuk, M S Ruhoff","doi":"10.1101/gr.4.5.299","DOIUrl":"https://doi.org/10.1101/gr.4.5.299","url":null,"abstract":"","PeriodicalId":77315,"journal":{"name":"PCR methods and applications","volume":"4 5","pages":"299-302"},"PeriodicalIF":0.0,"publicationDate":"1995-04-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"18586920","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}
{"title":"Amplification of the total coding sequence of the NF1 gene from peripheral blood lymphocyte RNA.","authors":"M H Shen, P S Harper, M Upadhyaya","doi":"10.1101/gr.4.5.311","DOIUrl":"https://doi.org/10.1101/gr.4.5.311","url":null,"abstract":"","PeriodicalId":77315,"journal":{"name":"PCR methods and applications","volume":"4 5","pages":"311-3"},"PeriodicalIF":0.0,"publicationDate":"1995-04-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"18586924","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}
Many groups have now published data based on the in situ detection of PCR-amplified DNA and cDNA. As with standard in situ hybridization or PCR, variables that can affect in situ PCR results include type of fixative and time of fixation, protease digestion, and the composition of the amplifying solution and oligoprobe cocktail. Investigators new to the field of in situ PCR should first try direct incorporation of the reporter molecule into paraffin-embedded tissue sections. Although nonspecific DNA synthesis is generated under these conditions, one can develop the confidence of synthesizing DNA inside the nucleus and appreciate the importance of protease digestion time to successful RT in situ PCR. It is an arguable statement that the in situ detection of PCR-amplified DNA and cDNA will have a very strong impact on many diverse fields, such as oncogenesis, embryology, RNA trafficking, and detection of viral diseases, as it already has on our understanding of the pathogenesis of HIV-1 infection.
{"title":"In situ PCR: protocols and applications.","authors":"G J Nuovo","doi":"10.1101/gr.4.4.s151","DOIUrl":"https://doi.org/10.1101/gr.4.4.s151","url":null,"abstract":"<p><p>Many groups have now published data based on the in situ detection of PCR-amplified DNA and cDNA. As with standard in situ hybridization or PCR, variables that can affect in situ PCR results include type of fixative and time of fixation, protease digestion, and the composition of the amplifying solution and oligoprobe cocktail. Investigators new to the field of in situ PCR should first try direct incorporation of the reporter molecule into paraffin-embedded tissue sections. Although nonspecific DNA synthesis is generated under these conditions, one can develop the confidence of synthesizing DNA inside the nucleus and appreciate the importance of protease digestion time to successful RT in situ PCR. It is an arguable statement that the in situ detection of PCR-amplified DNA and cDNA will have a very strong impact on many diverse fields, such as oncogenesis, embryology, RNA trafficking, and detection of viral diseases, as it already has on our understanding of the pathogenesis of HIV-1 infection.</p>","PeriodicalId":77315,"journal":{"name":"PCR methods and applications","volume":"4 4","pages":"S151-67"},"PeriodicalIF":0.0,"publicationDate":"1995-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"19554682","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}
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