Cry3A transgenic, bark Cry3A transgenic, bark beetle resistant sprucebeetle resistant spruce

NAZV project NAZV project QH71290QH712902007-20112007-2011

Spruce bark beetle (Ips typographus)

Bark beetle galleries

Bacillus thuringiensiscrystal protein (Cry, δ-endoTOXin)

Agrobacterium interaction with the plant cell

• Spruce is difficult to transformSpruce is difficult to transform

• No known Bt toxin is toxic enough to bark No known Bt toxin is toxic enough to bark beetlebeetle

Major problems:Major problems:

Spruce callus cultures

Somatic embryogenesis

induction conversion cultivation

Modified Cry3A genes

Cry3A orig

Synt TOX-1 (602 aa)-codons Picea optimized

Ndel/sTOX-1

Cdel/sTOX-1 (602 aa)

Cdel/Ndel/sTOX-1 GUS

Refined modification of domain II

Expression vector

PAGE analysis of modified Cry3A expression

WESTERN detection of modified Cry3A proteins

Toxicity tests on live beetles

K o n e c p r e s e n t a c e