Studies on Development of Efficient Transgenic System for Alien Gene of Insecticidal Protein in Sugarbeet

1999 Fiscal Year Final Research Report Summary

• Project/Area Number: 09460001
• Research Category: Grant-in-Aid for Scientific Research (B)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Breeding science
• Research Institution: HOKKAIDO UNIVERSITY
• Principal Investigator: SHIMAMOTO Yoshiya Hokkaido Univ., Grad. School of Agri., Pro., 大学院・農学研究科, 教授 (00001438)
• Co-Investigator(Kenkyū-buntansha): KANAZAWA Akira Hokkaido Univ., Grad. School of Agri., Inst., 大学院・農学研究科, 助手 (30281794) ; ASANO Shinichiro Hokkaido Univ., Grad. School of Agri., Asso. Pro., 大学院・農学研究科, 助教授 (60222585) ; SANO Yoshio Hokkaido Univ., Grad. School of Agri., Pro., 大学院・農学研究科, 教授 (70109528)
• Project Period (FY): 1997 – 1999
• Keywords: sugarbeet / cabbage armyworm / cry gene / ICP / Agrobacterium / bioassay / bombardment / chloroplast

Research Abstract

1)Development of chloroplast transgenic system in sugarbeet
We have progressed to develop a chloroplast transgenic system in sugarbeet in which is introduced cry genes to chloroplast genome and produce efficiently insecticidal crystal protein (ICP) in chloroplast. Plasmid carrying selective factor (aadA), marker factor (GUS), promoter (rrn) and terminator (rps 16) was incorporated into disc tissue of shoot of sugarbeet by microprojectile bombardment. In the tissue culture regeneration plant has not achieved. Repeatedly plasmid was constructed and incorporated into the sugarbeet.
2)Development of concentration system of protein upon organelle in sugarbeet
We have progressed to develop a concentration system of toxic protein upon organelle in sugarbeet. Base sequence coding transit peptide to chloroplast was cloned from tobacco plant and was linked with position of N terminal in GUS gene. Thus the plasmid was constructed and introduced into sugarbeet by Agrobacterium method.
3)Expression of cry gene in transformant sugarbeet
Several transformants carrying cryIA(b) were detected by PCR southern, ICP analysis and PCR southern blotting in half of plants regenerating after procedure of Agrobacterium method. Several leaves of these transformants were supplied for cabbage armyworm with the third larva age. Larva of cabbage armyworm was able to grow normally on leaves with non-cry gene and not to grow on several leaves of transformant.
4)Screening for efficient cry gene to produce insecticidal protein
Screening for efficient cry gene to produce insecticidal protein was conducted for cabbage armyworm. CryIC gene has more insecticidal ability than cryIA(b) in larva of cabbage armyworm.