Genotyping of insertion/deletion polymorphisms using locked nucleic acids for human identification

• Project/Area Number: 24590847
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Research Field: Legal medicine
• Research Institution: Asahikawa Medical College
• Principal Investigator: ASARI Masaru 旭川医科大学, 医学部, 准教授 (40360979)
• Co-Investigator(Kenkyū-buntansha): OMURA Tomohiro 京都大学, 医学(系)研究科(研究院), 助教 (00430935) ; OKA Kumiko 旭川医科大学, 医学部, 助教 (60571891) ; 大村 友博 京都大学, 医学(系)研究科(研究院), 助教 (00439035)
• Project Period (FY): 2012-04-01 – 2015-03-31
• Project Status: Completed (Fiscal Year 2014)
• Budget Amount: ¥5,460,000 (Direct Cost: ¥4,200,000、Indirect Cost: ¥1,260,000); Fiscal Year 2014: ¥1,300,000 (Direct Cost: ¥1,000,000、Indirect Cost: ¥300,000); Fiscal Year 2013: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000); Fiscal Year 2012: ¥2,730,000 (Direct Cost: ¥2,100,000、Indirect Cost: ¥630,000)
• Keywords: 個人識別 / 挿入欠失多型 / 人工核酸LNA / ユニバーサルプライマー / 人工核酸ＬＮＡ / PCR / 蛍光標識

Outline of Final Research Achievements

We analyzed the effects of locked nucleic acids (LNAs) for the efficiency of fluorescent labeling of PCR products. When the 24-base universal sequence with a higher Tm value was used, the introduction of a few LNAs significantly improved the efficiency, compared to the results from the 18-base universal sequence. We also selected 37 autosomal insertion/deletion (Indel) polymorphisms, and calculated the frequencies of alleles from 100 Japanese individuals. The matching probability was 2×10(-15), and the combination of selected Indels would be useful for individual discrimination. In multiplex Indel genotyping, fluorescent universal primers with LNAs produced higher signals than non-LNA fluorescent universal primers, and the increasing effects were identified especially in the low concentration (0.1 μM) of these primers.

Research Products

• [Journal Article] Rapid genotyping of 25 autosomal STRs in a Japanese population using fluorescent universal primers containing locked nucleic acids (2015)
  • Author(s): M. Asari, K. Okuda, D. Yajima, C. Maseda, C. Hoshina, T. Omura, H. Shiono, K. Matsubara, K. Shimizu.
  • Journal Title: Journal of Forensic and Legal Medicine Volume: 31 Pages: 36-41
  • DOI: 10.1016/j.jflm.2015.01.004
  • Peer Reviewed / Acknowledgement Compliant
• [Journal Article] Genotyping of 38 insertion/deletion polymorphisms for human identification using universal fluorescent PCR (2014)
  • Author(s): K. Oka, M. Asari, T. Omura, M. Yoshida, C. Maseda, D. Yajima, K. Matsubara, H. Shiono, M. Matsuda, K. Shimizu.
  • Journal Title: Molecular and Cellular Probes Volume: 28 Issue: 1 Pages: 13-18
  • DOI: 10.1016/j.mcp.2013.09.002
  • Peer Reviewed
• [Journal Article] 多座位挿入欠失多型解析に基づく個人識別法の開発 (2013)
  • Author(s): 岡 久美子, 浅利 優, 間瀬田 千香暁, 塩野 寛, 清水 惠子, 松田 光悦, 大村 友博, 松原 和夫, 吉田 将亜.
  • Journal Title: DNA多型 Volume: 21 Pages: 214-217
• [Presentation] Fast PCR-based genotyping of 25 autosomal short tandem repeats and amelogenin using fluorescent universal primers (2014)
  • Author(s): M. Asari, K. Oka, C. Hoshina, S. Isozaki, K. Okuda, D. Yajima, C. Maseda, T. Omura, K. Matsubara, H. Shiono, K. Shimizu.
  • Organizer: 9th International Symposium on Advances in Legal Medicine (ISALM)
  • Place of Presentation: Fukuoka
  • Year and Date: 2014-06-18
• [Presentation] Genotyping of 38 insertion/deletion polymorphisms for huma identification using universal fluorescent PCR (2014)
  • Author(s): K. Oka, M. Asari, T. Omura, S. Isozaki, M. Yoshida, C. Maseda, H. Shiono, M. Matsuda, K. Shimizu.
  • Organizer: 9th International Symposium on Advances in Legal Medicine (ISALM)
  • Place of Presentation: Fukuoka
  • Year and Date: 2014-06-17
• [Presentation] A new multiplex system for 38 insertion-deletion polymorphism analysis in Japanese individuals
  • Author(s): K. Oka, M. Asari, T. Omura, S. Isozaki, C. Ichimaru, M. Yoshida, K. Matsubara, H. Shiono, M. Matsuda, K. Shimizu.
  • Organizer: 日本法医学会
  • Place of Presentation: 札幌