Imaging of individual transcripts by amplification-based single-molecule fluorescence in situ hybridization
•A novel method termed asmFISH for in situ imaging of individual RNA molecules has been developed.•AsmFISH provides improved detection efficiency and specificity.•AsmFISH can be used for in situ SNP genotyping.•AsmFISH is applicable to fixed cell samples as well as fresh frozen or FFPE tissue sectio...
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| Veröffentlicht in: | New biotechnology 2021-03, Vol.61, p.116-123 |
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| container_title | New biotechnology |
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| creator | Lin, Chen Jiang, Meng Liu, Ling Chen, Xiaoyuan Zhao, Yuancun Chen, Lu Hong, Yujuan Wang, Xin Hong, Chengye Yao, Xihu Ke, Rongqin |
| description | •A novel method termed asmFISH for in situ imaging of individual RNA molecules has been developed.•AsmFISH provides improved detection efficiency and specificity.•AsmFISH can be used for in situ SNP genotyping.•AsmFISH is applicable to fixed cell samples as well as fresh frozen or FFPE tissue sections.
An amplification-based single-molecule fluorescence in situ hybridization (asmFISH) assay is introduced that exploits improved probe design for highly specific imaging of individual transcripts in fixed cells and tissues. In this method, a pair of DNA ligation probes are ligated on RNA templates upon specific hybridization, followed by probe circularization based on enzymatic DNA ligation and rolling circle amplification for signal boosting. The method is more efficient and specific than the padlock probe assay for detection of the same RNA molecules and discrimination of single nucleotide polymorphisms. Moreover, asmFISH is a versatile method which can be applied not only to cultured cells, but also to fresh frozen and formalin-fixed, paraffin-embedded tissue sections. |
| doi_str_mv | 10.1016/j.nbt.2020.12.001 |
| format | Article |
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An amplification-based single-molecule fluorescence in situ hybridization (asmFISH) assay is introduced that exploits improved probe design for highly specific imaging of individual transcripts in fixed cells and tissues. In this method, a pair of DNA ligation probes are ligated on RNA templates upon specific hybridization, followed by probe circularization based on enzymatic DNA ligation and rolling circle amplification for signal boosting. The method is more efficient and specific than the padlock probe assay for detection of the same RNA molecules and discrimination of single nucleotide polymorphisms. Moreover, asmFISH is a versatile method which can be applied not only to cultured cells, but also to fresh frozen and formalin-fixed, paraffin-embedded tissue sections.</description><identifier>ISSN: 1871-6784</identifier><identifier>EISSN: 1876-4347</identifier><identifier>DOI: 10.1016/j.nbt.2020.12.001</identifier><identifier>PMID: 33301924</identifier><language>eng</language><publisher>Netherlands: Elsevier B.V</publisher><subject>Amplification ; biotechnology ; Deoxyribonucleic acid ; DNA ; DNA ligation ; DNA probes ; Fluorescence ; Fluorescence in situ hybridization ; Gene expression ; Hybridization ; Nucleotides ; Padlock probes ; Paraffin ; Paraffins ; Ribonucleic acid ; RNA ; RNA probes ; Rolling circle amplification ; Single-nucleotide polymorphism</subject><ispartof>New biotechnology, 2021-03, Vol.61, p.116-123</ispartof><rights>2020 Elsevier B.V.</rights><rights>Copyright © 2020 Elsevier B.V. All rights reserved.</rights><rights>Copyright Elsevier Science Ltd. Mar 25, 2021</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c414t-c9f9787a12cbcc03fa82cd1fcbfe06b0a0cbc93030a4b5dea04672b9393338ce3</citedby><cites>FETCH-LOGICAL-c414t-c9f9787a12cbcc03fa82cd1fcbfe06b0a0cbc93030a4b5dea04672b9393338ce3</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktohtml>$$Uhttps://dx.doi.org/10.1016/j.nbt.2020.12.001$$EHTML$$P50$$Gelsevier$$H</linktohtml><link.rule.ids>314,778,782,3539,27907,27908,45978</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/33301924$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Lin, Chen</creatorcontrib><creatorcontrib>Jiang, Meng</creatorcontrib><creatorcontrib>Liu, Ling</creatorcontrib><creatorcontrib>Chen, Xiaoyuan</creatorcontrib><creatorcontrib>Zhao, Yuancun</creatorcontrib><creatorcontrib>Chen, Lu</creatorcontrib><creatorcontrib>Hong, Yujuan</creatorcontrib><creatorcontrib>Wang, Xin</creatorcontrib><creatorcontrib>Hong, Chengye</creatorcontrib><creatorcontrib>Yao, Xihu</creatorcontrib><creatorcontrib>Ke, Rongqin</creatorcontrib><title>Imaging of individual transcripts by amplification-based single-molecule fluorescence in situ hybridization</title><title>New biotechnology</title><addtitle>N Biotechnol</addtitle><description>•A novel method termed asmFISH for in situ imaging of individual RNA molecules has been developed.•AsmFISH provides improved detection efficiency and specificity.•AsmFISH can be used for in situ SNP genotyping.•AsmFISH is applicable to fixed cell samples as well as fresh frozen or FFPE tissue sections.
An amplification-based single-molecule fluorescence in situ hybridization (asmFISH) assay is introduced that exploits improved probe design for highly specific imaging of individual transcripts in fixed cells and tissues. In this method, a pair of DNA ligation probes are ligated on RNA templates upon specific hybridization, followed by probe circularization based on enzymatic DNA ligation and rolling circle amplification for signal boosting. The method is more efficient and specific than the padlock probe assay for detection of the same RNA molecules and discrimination of single nucleotide polymorphisms. Moreover, asmFISH is a versatile method which can be applied not only to cultured cells, but also to fresh frozen and formalin-fixed, paraffin-embedded tissue sections.</description><subject>Amplification</subject><subject>biotechnology</subject><subject>Deoxyribonucleic acid</subject><subject>DNA</subject><subject>DNA ligation</subject><subject>DNA probes</subject><subject>Fluorescence</subject><subject>Fluorescence in situ hybridization</subject><subject>Gene expression</subject><subject>Hybridization</subject><subject>Nucleotides</subject><subject>Padlock probes</subject><subject>Paraffin</subject><subject>Paraffins</subject><subject>Ribonucleic acid</subject><subject>RNA</subject><subject>RNA probes</subject><subject>Rolling circle amplification</subject><subject>Single-nucleotide polymorphism</subject><issn>1871-6784</issn><issn>1876-4347</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2021</creationdate><recordtype>article</recordtype><recordid>eNqFkT1vFDEQhi0EIiHwA2jQSjQ0e4w_br0WFYoCRIpEA7Xlj3Hw4V0f9m6k49fjywUKCqg89jzvK8-8hLyksKFAh7e7zWyXDQPW7mwDQB-RczrKoRdcyMf3Ne0HOYoz8qzWHcBA1UCfkjPOOVDFxDn5fj2Z2zjfdjl0cfbxLvrVpG4pZq6uxP1SO3vozLRPMURnlpjn3pqKvqtNlbCfckK3JuxCWnPB6nB22Kxaf1m7bwdboo8_74XPyZNgUsUXD-cF-frh6svlp_7m88fry_c3vRNULL1TQclRGsqcdQ54MCNzngZnA8JgwUB7Vxw4GGG3Hg2IQTKruGpjjQ75BXlz8t2X_GPFuugptn-lZGbMa9VMjVshQSr2f1RIRmHLtqqhr_9Cd3ktcxukUYoqyUZ2NKQnypVca8Gg9yVOphw0BX0MTe90C00fQ9OU6RZa07x6cF7thP6P4ndKDXh3ArBt7S5i0dXF4559LOgW7XP8h_0vwSGpAw</recordid><startdate>20210325</startdate><enddate>20210325</enddate><creator>Lin, Chen</creator><creator>Jiang, Meng</creator><creator>Liu, Ling</creator><creator>Chen, Xiaoyuan</creator><creator>Zhao, Yuancun</creator><creator>Chen, Lu</creator><creator>Hong, Yujuan</creator><creator>Wang, Xin</creator><creator>Hong, Chengye</creator><creator>Yao, Xihu</creator><creator>Ke, Rongqin</creator><general>Elsevier B.V</general><general>Elsevier Science Ltd</general><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7QO</scope><scope>8FD</scope><scope>FR3</scope><scope>P64</scope><scope>7X8</scope><scope>7S9</scope><scope>L.6</scope></search><sort><creationdate>20210325</creationdate><title>Imaging of individual transcripts by amplification-based single-molecule fluorescence in situ hybridization</title><author>Lin, Chen ; Jiang, Meng ; Liu, Ling ; Chen, Xiaoyuan ; Zhao, Yuancun ; Chen, Lu ; Hong, Yujuan ; Wang, Xin ; Hong, Chengye ; Yao, Xihu ; Ke, Rongqin</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c414t-c9f9787a12cbcc03fa82cd1fcbfe06b0a0cbc93030a4b5dea04672b9393338ce3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2021</creationdate><topic>Amplification</topic><topic>biotechnology</topic><topic>Deoxyribonucleic acid</topic><topic>DNA</topic><topic>DNA ligation</topic><topic>DNA probes</topic><topic>Fluorescence</topic><topic>Fluorescence in situ hybridization</topic><topic>Gene expression</topic><topic>Hybridization</topic><topic>Nucleotides</topic><topic>Padlock probes</topic><topic>Paraffin</topic><topic>Paraffins</topic><topic>Ribonucleic acid</topic><topic>RNA</topic><topic>RNA probes</topic><topic>Rolling circle amplification</topic><topic>Single-nucleotide polymorphism</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Lin, Chen</creatorcontrib><creatorcontrib>Jiang, Meng</creatorcontrib><creatorcontrib>Liu, Ling</creatorcontrib><creatorcontrib>Chen, Xiaoyuan</creatorcontrib><creatorcontrib>Zhao, Yuancun</creatorcontrib><creatorcontrib>Chen, Lu</creatorcontrib><creatorcontrib>Hong, Yujuan</creatorcontrib><creatorcontrib>Wang, Xin</creatorcontrib><creatorcontrib>Hong, Chengye</creatorcontrib><creatorcontrib>Yao, Xihu</creatorcontrib><creatorcontrib>Ke, Rongqin</creatorcontrib><collection>PubMed</collection><collection>CrossRef</collection><collection>Biotechnology Research Abstracts</collection><collection>Technology Research Database</collection><collection>Engineering Research Database</collection><collection>Biotechnology and BioEngineering Abstracts</collection><collection>MEDLINE - Academic</collection><collection>AGRICOLA</collection><collection>AGRICOLA - Academic</collection><jtitle>New biotechnology</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Lin, Chen</au><au>Jiang, Meng</au><au>Liu, Ling</au><au>Chen, Xiaoyuan</au><au>Zhao, Yuancun</au><au>Chen, Lu</au><au>Hong, Yujuan</au><au>Wang, Xin</au><au>Hong, Chengye</au><au>Yao, Xihu</au><au>Ke, Rongqin</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Imaging of individual transcripts by amplification-based single-molecule fluorescence in situ hybridization</atitle><jtitle>New biotechnology</jtitle><addtitle>N Biotechnol</addtitle><date>2021-03-25</date><risdate>2021</risdate><volume>61</volume><spage>116</spage><epage>123</epage><pages>116-123</pages><issn>1871-6784</issn><eissn>1876-4347</eissn><abstract>•A novel method termed asmFISH for in situ imaging of individual RNA molecules has been developed.•AsmFISH provides improved detection efficiency and specificity.•AsmFISH can be used for in situ SNP genotyping.•AsmFISH is applicable to fixed cell samples as well as fresh frozen or FFPE tissue sections.
An amplification-based single-molecule fluorescence in situ hybridization (asmFISH) assay is introduced that exploits improved probe design for highly specific imaging of individual transcripts in fixed cells and tissues. In this method, a pair of DNA ligation probes are ligated on RNA templates upon specific hybridization, followed by probe circularization based on enzymatic DNA ligation and rolling circle amplification for signal boosting. The method is more efficient and specific than the padlock probe assay for detection of the same RNA molecules and discrimination of single nucleotide polymorphisms. Moreover, asmFISH is a versatile method which can be applied not only to cultured cells, but also to fresh frozen and formalin-fixed, paraffin-embedded tissue sections.</abstract><cop>Netherlands</cop><pub>Elsevier B.V</pub><pmid>33301924</pmid><doi>10.1016/j.nbt.2020.12.001</doi><tpages>8</tpages></addata></record> |
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| source | ScienceDirect Journals (5 years ago - present) |
| subjects | Amplification biotechnology Deoxyribonucleic acid DNA DNA ligation DNA probes Fluorescence Fluorescence in situ hybridization Gene expression Hybridization Nucleotides Padlock probes Paraffin Paraffins Ribonucleic acid RNA RNA probes Rolling circle amplification Single-nucleotide polymorphism |
| title | Imaging of individual transcripts by amplification-based single-molecule fluorescence in situ hybridization |
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