Analysis of mechanisms of regeneration-bud formation in the cricket using novel genetic engineering method

2017 Fiscal Year Final Research Report

• Project/Area Number: 16K21199
• Research Category: Grant-in-Aid for Young Scientists (B)
• Allocation Type: Multi-year Fund
• Research Field: Integrative animal science; Insect science
• Research Institution: Tokushima Agriculture, Forestry, and Fisheries Technology Support Center
• Principal Investigator: WATANABE TAKAHITO 徳島県立農林水産総合技術支援センター(試験研究部), 徳島県立農林水産総合技術支援センター(資源環境研究課), 研究員 (30709481)
• Project Period (FY): 2016-04-01 – 2018-03-31
• Keywords: ゲノム編集 / フタホシコオロギ / ノックイン / RNA-seq

Outline of Final Research Achievements

Using genome editing technology, knock-in vector containing eGFP gene under cricket minimal promoter was introduced into Hox gene loci in the cricket genome. eGFP fluorescent was observed as expected pattern. We succeeded to establish the method for generating enhancer trap line in the cricket. For introducing correctly knock-in direction and junction, knock-in method via MMEJ pathway was applied in the cricket. eGFP gene was correctly introduced into stop codon loci in target gene via MMEJ mediated knock-in, and eGFP expression was observed. In addition, RNA-seq analysis was performed using PacBioRSII. Extracted mRNA from cricket embryo and nymph was fractionated into four, and each fraction was sequenced in 4 cells. As a result, 760,000 reads and 2.25 Gb sequence was obtained in total.

Free Research Field

昆虫科学