Early Development of Cloned Bovine Embryos Produced from Oocytes Enucleated by Fluorescence Metaphase II Imaging Using a Conventional Halogen-Lamp Microscope
Enucleation of a recipient oocyte is one of the key processes in the procedure of somatic cell nuclear transfer (SCNT). However, especially in bovine species, lipid droplets spreading in the ooplasm hamper identification and enucleation of metaphase II (MII) chromosomes, and thereby the success rate...
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| Veröffentlicht in: | Cellular reprogramming 2015-04, Vol.17 (2), p.106-114 |
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| container_title | Cellular reprogramming |
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| creator | Iwamoto, Daisaku Yamagata, Kazuo Kishi, Masao Hayashi-Takanaka, Yoko Kimura, Hiroshi Wakayama, Teruhiko Saeki, Kazuhiro |
| description | Enucleation of a recipient oocyte is one of the key processes in the procedure of somatic cell nuclear transfer (SCNT). However, especially in bovine species, lipid droplets spreading in the ooplasm hamper identification and enucleation of metaphase II (MII) chromosomes, and thereby the success rate of the cloning remains low. In this study we used a new experimental system that enables fluorescent observation of chromosomes in living oocytes without any damage. We succeeded in visualizing and removing the MII chromosome in matured bovine oocytes. This experimental system consists of injecting fluorescence-labeled antibody conjugates that bind to chromosomes and fluorescent observation using a conventional halogen-lamp microscope. The cleavage rates and blastocyst rates of bovine embryos following in vitro fertilization (IVF) decreased as the concentration of the antibody increased (p0.05). For the production of SCNT embryos, the new system can be used as a reliable predictor of the location of metaphase plates in opaque oocytes, such as those in ruminant animals. |
| doi_str_mv | 10.1089/cell.2014.0086 |
| format | Article |
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However, especially in bovine species, lipid droplets spreading in the ooplasm hamper identification and enucleation of metaphase II (MII) chromosomes, and thereby the success rate of the cloning remains low. In this study we used a new experimental system that enables fluorescent observation of chromosomes in living oocytes without any damage. We succeeded in visualizing and removing the MII chromosome in matured bovine oocytes. This experimental system consists of injecting fluorescence-labeled antibody conjugates that bind to chromosomes and fluorescent observation using a conventional halogen-lamp microscope. The cleavage rates and blastocyst rates of bovine embryos following in vitro fertilization (IVF) decreased as the concentration of the antibody increased (p<0.05). The enucleation rate of the conventional method (blind enucleation) was 86%, whereas all oocytes injected with the antibody conjugates were enucleated successfully. Fusion rates and developmental rates of SCNT embryos produced with the enucleated oocytes were the same as those of the blind enucleation group (p>0.05). For the production of SCNT embryos, the new system can be used as a reliable predictor of the location of metaphase plates in opaque oocytes, such as those in ruminant animals.</description><identifier>ISSN: 2152-4998</identifier><identifier>ISSN: 2152-4971</identifier><identifier>EISSN: 2152-4998</identifier><identifier>DOI: 10.1089/cell.2014.0086</identifier><identifier>PMID: 25826723</identifier><language>eng</language><publisher>United States: Mary Ann Liebert, Inc</publisher><subject>Animals ; Blastocyst - cytology ; Cattle ; Chromosomes ; Cloning, Organism - methods ; Embryonic Development ; Fertilization in Vitro - veterinary ; Fluorescence ; Immunoconjugates - chemistry ; Male ; Metaphase ; Nuclear Transfer Techniques - veterinary ; Oocytes - cytology ; Original Research ; Phycoerythrin - chemistry</subject><ispartof>Cellular reprogramming, 2015-04, Vol.17 (2), p.106-114</ispartof><rights>Copyright 2015, Mary Ann Liebert, Inc. 2015</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c524t-b327c4a64bae45c55b33387dea263ba943cacec52969026d4c0d377efdcfd4ae3</citedby><cites>FETCH-LOGICAL-c524t-b327c4a64bae45c55b33387dea263ba943cacec52969026d4c0d377efdcfd4ae3</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>230,314,780,784,885,27924,27925</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/25826723$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Iwamoto, Daisaku</creatorcontrib><creatorcontrib>Yamagata, Kazuo</creatorcontrib><creatorcontrib>Kishi, Masao</creatorcontrib><creatorcontrib>Hayashi-Takanaka, Yoko</creatorcontrib><creatorcontrib>Kimura, Hiroshi</creatorcontrib><creatorcontrib>Wakayama, Teruhiko</creatorcontrib><creatorcontrib>Saeki, Kazuhiro</creatorcontrib><title>Early Development of Cloned Bovine Embryos Produced from Oocytes Enucleated by Fluorescence Metaphase II Imaging Using a Conventional Halogen-Lamp Microscope</title><title>Cellular reprogramming</title><addtitle>Cell Reprogram</addtitle><description>Enucleation of a recipient oocyte is one of the key processes in the procedure of somatic cell nuclear transfer (SCNT). However, especially in bovine species, lipid droplets spreading in the ooplasm hamper identification and enucleation of metaphase II (MII) chromosomes, and thereby the success rate of the cloning remains low. In this study we used a new experimental system that enables fluorescent observation of chromosomes in living oocytes without any damage. We succeeded in visualizing and removing the MII chromosome in matured bovine oocytes. This experimental system consists of injecting fluorescence-labeled antibody conjugates that bind to chromosomes and fluorescent observation using a conventional halogen-lamp microscope. The cleavage rates and blastocyst rates of bovine embryos following in vitro fertilization (IVF) decreased as the concentration of the antibody increased (p<0.05). The enucleation rate of the conventional method (blind enucleation) was 86%, whereas all oocytes injected with the antibody conjugates were enucleated successfully. Fusion rates and developmental rates of SCNT embryos produced with the enucleated oocytes were the same as those of the blind enucleation group (p>0.05). For the production of SCNT embryos, the new system can be used as a reliable predictor of the location of metaphase plates in opaque oocytes, such as those in ruminant animals.</description><subject>Animals</subject><subject>Blastocyst - cytology</subject><subject>Cattle</subject><subject>Chromosomes</subject><subject>Cloning, Organism - methods</subject><subject>Embryonic Development</subject><subject>Fertilization in Vitro - veterinary</subject><subject>Fluorescence</subject><subject>Immunoconjugates - chemistry</subject><subject>Male</subject><subject>Metaphase</subject><subject>Nuclear Transfer Techniques - veterinary</subject><subject>Oocytes - cytology</subject><subject>Original Research</subject><subject>Phycoerythrin - chemistry</subject><issn>2152-4998</issn><issn>2152-4971</issn><issn>2152-4998</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2015</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNpVkcFu1DAQhiMEolXplSP4yCWLYztOckGCZUtX2qpIsGdr4kzSIMcOdrJSHoZ3xdGWqliyxxp__8xYf5K8zegmo2X1UaMxG0YzsaG0lC-SS5blLBVVVb58dr9IrkP4RePiPMrk6-SC5SWTBeOXyZ8deLOQr3hC48YB7URcS7bGWWzIF3fqLZLdUPvFBfLdu2bWMd96N5B7p5cJA9nZWRuEKebrhdyY2XkMGq1GcocTjA8QkOz3ZD9A19uOHMN6Atk6e4rtemfBkFswrkObHmAYyV2vvQvajfgmedWCCXj9GK-S483u5_Y2Pdx_228_H1KdMzGlNWeFFiBFDShynec157wsGgQmeQ2V4Bo0RraSFWWyEZo2vCiwbXTbCEB-lXw61x3nesAmTj95MGr0_QB-UQ569f-L7R9U505K8KIsizIW-PBYwLvfM4ZJDX1Y7QGLbg4qk7ISOc0qFtHNGV0_GTy2T20yqlZb1apTq61qtTUK3j0f7gn_Z2IE3p-BFpyCzvdBHX9EvaRxFzTL-V9CVKvc</recordid><startdate>20150401</startdate><enddate>20150401</enddate><creator>Iwamoto, Daisaku</creator><creator>Yamagata, Kazuo</creator><creator>Kishi, Masao</creator><creator>Hayashi-Takanaka, Yoko</creator><creator>Kimura, Hiroshi</creator><creator>Wakayama, Teruhiko</creator><creator>Saeki, Kazuhiro</creator><general>Mary Ann Liebert, Inc</general><scope>FBQ</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7X8</scope><scope>5PM</scope></search><sort><creationdate>20150401</creationdate><title>Early Development of Cloned Bovine Embryos Produced from Oocytes Enucleated by Fluorescence Metaphase II Imaging Using a Conventional Halogen-Lamp Microscope</title><author>Iwamoto, Daisaku ; Yamagata, Kazuo ; Kishi, Masao ; Hayashi-Takanaka, Yoko ; Kimura, Hiroshi ; Wakayama, Teruhiko ; Saeki, Kazuhiro</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c524t-b327c4a64bae45c55b33387dea263ba943cacec52969026d4c0d377efdcfd4ae3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2015</creationdate><topic>Animals</topic><topic>Blastocyst - cytology</topic><topic>Cattle</topic><topic>Chromosomes</topic><topic>Cloning, Organism - methods</topic><topic>Embryonic Development</topic><topic>Fertilization in Vitro - veterinary</topic><topic>Fluorescence</topic><topic>Immunoconjugates - chemistry</topic><topic>Male</topic><topic>Metaphase</topic><topic>Nuclear Transfer Techniques - veterinary</topic><topic>Oocytes - cytology</topic><topic>Original Research</topic><topic>Phycoerythrin - chemistry</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Iwamoto, Daisaku</creatorcontrib><creatorcontrib>Yamagata, Kazuo</creatorcontrib><creatorcontrib>Kishi, Masao</creatorcontrib><creatorcontrib>Hayashi-Takanaka, Yoko</creatorcontrib><creatorcontrib>Kimura, Hiroshi</creatorcontrib><creatorcontrib>Wakayama, Teruhiko</creatorcontrib><creatorcontrib>Saeki, Kazuhiro</creatorcontrib><collection>AGRIS</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>MEDLINE - Academic</collection><collection>PubMed Central (Full Participant titles)</collection><jtitle>Cellular reprogramming</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Iwamoto, Daisaku</au><au>Yamagata, Kazuo</au><au>Kishi, Masao</au><au>Hayashi-Takanaka, Yoko</au><au>Kimura, Hiroshi</au><au>Wakayama, Teruhiko</au><au>Saeki, Kazuhiro</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Early Development of Cloned Bovine Embryos Produced from Oocytes Enucleated by Fluorescence Metaphase II Imaging Using a Conventional Halogen-Lamp Microscope</atitle><jtitle>Cellular reprogramming</jtitle><addtitle>Cell Reprogram</addtitle><date>2015-04-01</date><risdate>2015</risdate><volume>17</volume><issue>2</issue><spage>106</spage><epage>114</epage><pages>106-114</pages><issn>2152-4998</issn><issn>2152-4971</issn><eissn>2152-4998</eissn><abstract>Enucleation of a recipient oocyte is one of the key processes in the procedure of somatic cell nuclear transfer (SCNT). However, especially in bovine species, lipid droplets spreading in the ooplasm hamper identification and enucleation of metaphase II (MII) chromosomes, and thereby the success rate of the cloning remains low. In this study we used a new experimental system that enables fluorescent observation of chromosomes in living oocytes without any damage. We succeeded in visualizing and removing the MII chromosome in matured bovine oocytes. This experimental system consists of injecting fluorescence-labeled antibody conjugates that bind to chromosomes and fluorescent observation using a conventional halogen-lamp microscope. The cleavage rates and blastocyst rates of bovine embryos following in vitro fertilization (IVF) decreased as the concentration of the antibody increased (p<0.05). The enucleation rate of the conventional method (blind enucleation) was 86%, whereas all oocytes injected with the antibody conjugates were enucleated successfully. Fusion rates and developmental rates of SCNT embryos produced with the enucleated oocytes were the same as those of the blind enucleation group (p>0.05). For the production of SCNT embryos, the new system can be used as a reliable predictor of the location of metaphase plates in opaque oocytes, such as those in ruminant animals.</abstract><cop>United States</cop><pub>Mary Ann Liebert, Inc</pub><pmid>25826723</pmid><doi>10.1089/cell.2014.0086</doi><tpages>9</tpages><oa>free_for_read</oa></addata></record> |
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| subjects | Animals Blastocyst - cytology Cattle Chromosomes Cloning, Organism - methods Embryonic Development Fertilization in Vitro - veterinary Fluorescence Immunoconjugates - chemistry Male Metaphase Nuclear Transfer Techniques - veterinary Oocytes - cytology Original Research Phycoerythrin - chemistry |
| title | Early Development of Cloned Bovine Embryos Produced from Oocytes Enucleated by Fluorescence Metaphase II Imaging Using a Conventional Halogen-Lamp Microscope |
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