Rapid clonig and transfer of a huge genome DNA using human artificial chromosome

2017 Fiscal Year Final Research Report

• Project/Area Number: 15K06927
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Research Field: System genome science
• Research Institution: Kazusa DNA Research Institute
• Principal Investigator: Hasegawa Yoshinori 公益財団法人かずさDNA研究所, バイオ研究開発部, チーム長 (30387683)
• Project Period (FY): 2015-04-01 – 2018-03-31
• Keywords: 巨大ゲノム / クローニング / ヒト人工染色体 / 部位特異的組換え / Vlox / Slox

Outline of Final Research Achievements

It was increased an insertion efficiency of VLox and SLox sites, which are used for site specific recombination, on both sides of the target gene (GOI) by optimizing the reaction conditions of a CRISPR/Cas9 system. However, the efficiency varied greatly among cell types. On the other hand, the efficiency of the GOI into the HAC vector was little difference between cell types, and also succeeded for GOI with a total DNA length of 600 kb. It was suceeded in tarnsferring HAC vector carrying GOI to other cells in all cell types tested . By selecting the host cell type, it was possible to complete the process up to cloning of the large GOI into the HAC vector, and transfer of the HAC vector carrying the GOI to the target cell in 2 months.

Free Research Field

細胞工学