Modification of piggyBac transposase gene on its sequence specificity

• Project/Area Number: 24658057
• Research Category: Grant-in-Aid for Challenging Exploratory Research
• Allocation Type: Multi-year Fund
• Research Field: Applied entomology
• Research Institution: National Institute of Agrobiological Sciences
• Principal Investigator: KOJIMA Katsura 独立行政法人農業生物資源研究所, 新機能素材研究開発ユニット, 主任研究員 (40370655)
• Project Period (FY): 2012-04-01 – 2015-03-31
• Project Status: Completed (Fiscal Year 2014)
• Budget Amount: ¥3,900,000 (Direct Cost: ¥3,000,000、Indirect Cost: ¥900,000); Fiscal Year 2014: ¥1,040,000 (Direct Cost: ¥800,000、Indirect Cost: ¥240,000); Fiscal Year 2013: ¥1,170,000 (Direct Cost: ¥900,000、Indirect Cost: ¥270,000); Fiscal Year 2012: ¥1,690,000 (Direct Cost: ¥1,300,000、Indirect Cost: ¥390,000)
• Keywords: 遺伝子組換えカイコ / piggyBac / DNA結合タンパク質 / TTAA / ZincFinger Domain / ZincFinger / カイコ

Outline of Final Research Achievements

To improve transposase activity of piggyBac transposase on its target sequence specificity, the gene encoding piggyBac transposase was modified by adding specific DNA binding domain on its N-terminus and its transposase activity was analyzed.
Transposase activity was analyzed by transfection and microinjection of modified piggyBac transposase in BmN cells and silkworm eggs, and inserted sequence were cloned along with surrounding genomic sequence. By sequencing of these cloned DNA fragments indicates (A) modified piggyBac retains its transposase activity, but (B) no specific insertion event were detected.