RBRC10465 C (3-6 months) Necessary documents for ordering:<ol><li>Approval form (<A HREF="https://mus.brc.riken.jp/ja/wp-content/uploads/form/form_6.docx">Japanese</A> / <A HREF="https://mus.brc.riken.jp/en/wp-content/uploads/form/form_d.docx">English</A>)</li><li>Order form (<A HREF="https://mus.brc.riken.jp/ja/wp-content/uploads/form/form_4.docx">Japanese</A> / <A HREF="https://mus.brc.riken.jp/en/wp-content/uploads/form/form_b.docx">English</A>)</li><li>Category I MTA: CRISPR/Cas9 genome edited bioresources (<A HREF="https://mus.brc.riken.jp/ja/wp-content/uploads/form/Broad_MTA_J.docx">Japanese</A> / <A HREF="https://mus.brc.riken.jp/en/wp-content/uploads/form/Broad_MTA_E.docx">English</A>)</li><li>The RECIPIENT of BIOLOGICAL RESOURCE must obtain a prior written consent on use of it from the DEVELOPER of the Venus using Approval form (<a HREF="https://mus.brc.riken.jp/en/wp-content/uploads/form/Form_V.docx">Form V (English)</a>) (Lab Contact: Laboratory for Cell Function Dynamics, RIKEN CBS: mta-cfds@ml.riken.jp).</li><li>Acceptance of responsibility for living modified organism (<A HREF="https://mus.brc.riken.jp/ja/wp-content/uploads/form/form_7.docx">Japanese</A> / <A HREF="https://mus.brc.riken.jp/en/wp-content/uploads/form/form_g.docx">English</A>)</li></ol> 条件を付加する。利用者は提供承諾書を用いて、事前に寄託者の承諾を得る。<br>研究成果の公表にあたって寄託者の指定する文献を引用する。後日指定<br>研究成果の公表にあたって謝辞の表明を必要とする。<br>利用者が本件リソースを使用して得られた研究成果に基づき特許等の申請、及び事業活動を行う場合は、寄託者と別途協議を行う。<br>利用者はApproval form (form V)を用いて、事前にVenusの開発者の許可を得る。<br>（連絡先：Laboratory for Cell Function Dynamics, RIKEN CBS: mta-cfds@ml.riken.jp) Oplophorus gracilirostris NanoLuc cDNA, synthetic FLAG (3xFLAG tag), Thoseaasigna virus 2A peptide (T2A), Aequorea victoria mVenus (variant of enhanced yellow fluorescent protein) cDNA, Bos taurus GH (Growth hormone polyA), mouse Amy1 genomic DNA <a href='https://brc.riken.jp/mus/pcr10465'>Genotyping protocol -PCR-</a> Amy1 Amy1 Reporter mouse strain which can monitor Amylase 1 (Amy1) secretion with the NanoLuc (Nluc) and Amy1 expressing cells with mVenus. A DNA construct, Nluc-3xFLAG-T2A-mVenus-BGHpA, was knocked in at downstream of the Amy gene promoter. C（3〜6か月） 井上直和・関亦正幸（福島県立医科大学）、関亦明子（山形大学）らによって２０１８年に独自開発し、以降、当研究室にて維持。B6D2F1マウス受精卵にCRISPR/Cas9ゲノム編集技術を用いてNanoLucルシフェラーゼ遺伝子とmVenus遺伝子をT2Aペプチドで連結したレポーター遺伝子をAmy1遺伝子座にノックインして作出。ファウンダーマウスをICRマウスと３世代交配し、その後、兄妹交配にてノックインホモマウスとして維持。野生型とマクロな解剖所見に違いなし。ホモマウスも正常に生育、交配できる。分泌液中にNanoLucルシフェラーゼ活性あり。唾液腺組織でmVenusを検出。 Fluorescent Proteins/lacZ System true STOCK Amy1<em1(luc/mVenus)Matam> a-アミラーゼ１遺伝子の分泌シグナルペプチドの下流にルシフェラーゼ遺伝子(NanoLuc)とmVenus遺伝子をT2Aで連結したNluc-3xFLAG-T2A-mVenus-BGHpAをノックインしたレポーター挿入マウス。これらレポーター遺伝子はa-アミラーゼ１遺伝子プロモーター制御下に発現する。そのため、ルシフェラーゼは細胞外に分泌されるため唾液分泌を定量化でき、mVenusは細胞質内に局在するためa-アミラーゼ１遺伝子発現細胞を検出できる。 Prior to requesting the BIOLOGICAL RESOURCE, the RECIPIENT must obtain approval from the DEPOSITOR using the Approval Form. In publishing the research results obtained by use of the BIOLOGICAL RESOURCE, a citation of the following literature(s) designated by the DEPOSITOR is requested. 後日指定In publishing the research results to be obtained by use of the BIOLOGICAL RESOURCE, an acknowledgment to the DEPOSITOR is requested. The RECIPIENT must contact the DEPOSITOR in the case of application for any patents or commercial use based on the results from the use of the BIOLOGICAL RESOURCE. The RECIPIENT of BIOLOGICAL RESOURCE must obtain a prior written consent on use of it from the DEVELOPER of the Venus using Approval form (form V) (Lab Contact: Laboratory for Cell Function Dynamics, RIKEN CBS: mta-cfds@ml.riken.jp). Developed by Naokazu Inoue and Masayuki Sekimata, Fukushima Medical University, and Akiko Sekimata, Yamagata University in 2018. This strain was generated by the CRISPR/Cas9 using fertilized eggs derived from B6D2F1. Mice were further crossed with ICR. Mixed genetic background.