Project Number: 6615-22000-021-16
Project Type: Nonfunded Cooperative Agreement

Start Date: Sep 27, 2007
End Date: Aug 31, 2008

Objective:
JcDNV somatic transformation vectors and piggyBac germ line vectors can be used to express high levels of proteins in insect cells. The proposed work will permit tests of modified vectors to optimize protein production and secretion of chimeric silk proteins incorporating segments of natural and synthetic silk genes from the silkmoth, Bombyx mori and the golden orb spider, Nauphoeta clavipes, and in insect cell culture and in vivo in Bombyx mori silk glands.

Approach:
The strategy is to combine the ability to efficiently somatically transform insect cells with the assessment of the importance of spider silk protein domains in producing large quantities of a high strength silk fiber. A chimeric silk protein gene called Spidrofibroin (SpF), which will combine the repetitive domains of the Nauphoeta clavipes dragline 1 protein with the N- and C- terminal domains of the Fibroin-H (Fib-H) protein from the silk moth Bombyx mori will be inserted into JcDNV somatic vectors that contain various insect promoters that result in high amounts of protein expression. These constructs will be used to transform insect cell lines and insect embryos to test levels of protein production in specific insect cell lines and tissues. The repetitive domains of dragline 1 produce the unique mechanical characteristics of the Nauphoeta clavipes dragline silk while the N- and C- terminal domains of Fib-H are essential for transport and assembly of Fib-H proteins into the silk fiber. Therefore when expressed from somatically transformed insect cell cultures or larvae, we expect that SpF will be transported and assembled into the silk fiber in a manner similar to that of Fib-H, but with the exceptional mechanical properties of spider silk. Confirmation of desirable characteristics will lead to germline transformation of Bombyx mori with high production chimeric silk genes to assess tensile strength of the chimeric protein.