Establishment of the efficient dsDNA delivery system into insect cells

2015 Fiscal Year Final Research Report

• Project/Area Number: 26660271
• Research Category: Grant-in-Aid for Challenging Exploratory Research
• Allocation Type: Multi-year Fund
• Research Field: Insect science
• Research Institution: Kyushu University
• Principal Investigator: LEE JAEMAN 九州大学, (連合)農学研究科(研究院), 助教 (50404083)
• Project Period (FY): 2014-04-01 – 2016-03-31
• Keywords: DNA導入法 / 線虫SID1 / dsDNA取込み / BmN4-SID1

Outline of Final Research Achievements

In order to establish the efficient dsDNA delivery system into insect cells, SID1 gene from Caenorhabditis elegans were introduced into various insect derived cell lines. Among the cells tested, silkworm derived PS140-SID1 cell exhibited the highest dsDNA uptake ability. Additional introduction of SID2, 3, 4, and 5 into BmN4-SID1 showed no effect on the dsDNA uptake. Also addition of divalent cations showed little increase of dsDNA uptake but the formation of hetero-duplex or triplex reduced the efficiency.
Then by using the fusion protein between lambda N protein with RNA binding activity and sso7d dsDNA binding protein, the RNA-dependent introduction of dsRNA was performed. As a result, the fusion protein more than 0.05 mM stimulated the dsDNA uptake in BmN4 cells. Also the attachment of sso7d or reverse LAMP2C peptide to the N-terminus of SID1 increased the dsDNA uptake.

Free Research Field

昆虫分子生物学