A procedure to enrich for auxotrophic and fermentation mutants of Agrobacterium tumefaciens is described. The method is based on the amplification of the killing power of carbenicillin by the addition of lysozyme. Isolation frequencies of some types of mutants are presented, with and without the application of the proposed procedure. The yield of mutants is usually enhanced a hundredfold per en...

An assay to determine the relative virulence of wild-type and mutant strains of Agrobacterium tumefaciens on leaves of Bryophyllum daigremontiana has been developed. This assay is reproducible, is easy to learn, is not time consuming, and requires little space. The relative virulence of cellulose-minus mutants of A. tumefaciens was investigated with this assay. Some of these mutants were unalte...

Bacteria use conjugation systems, a subfamily of the type IV secretion systems, to transfer DNA to recipient cells. Despite 50 years of research, the architecture and mechanism of action of the channel mediating DNA transfer across the bacterial cell envelope remains obscure. By use of a sensitive, quantifiable assay termed transfer DNA immunoprecipitation (TrIP), we identify contacts between a...

A binary vector is a standard tool in the transformation of higher plants mediated by Agrobacterium tumefaciens. It is composed of the borders of T-DNA, multiple cloning sites, replication functions for Escherichia coli and A. tumefaciens, selectable marker genes, reporter genes, and other accessory elements that can improve the efficiency of and/or give further capability to the system. A supe...

Agrobacterium tumefaciens-mediated transformation is an effective method for introducing genes into maize. In this chapter, we describe a detailed protocol for genetic transformation of the maize genotype Hi II. Our starting plant material is immature embryos cocultivated with an Agrobacterium strain carrying a standard binary vector. In addition to step-by-step laboratory transformation proced...

Crataeva religiosa G. Forst. var. nurvala (Buch.-Ham.) Hook. F. & Thomson (syn. Crataeva nurvala Buch.-Ham.), a fast growing medicinal tree species, has been successfully micropropagated under in vitro conditions. Direct multiple shoots were regenerated from nodal explants on Murashige and Skoog’s (MS) medium, solidified by agar and supplemented with 1 mg/L 6-benzyl amino purine (BAP) and 0.5 m...

Eggplant is highly responsive to various tissue culture techniques. Somatic embryogenesis and direct organogenesis are widely studied protocols in this crop, but potential of regeneration varies with genotype, explant and culture media supplemented with different combination and concentration of growth hormones. The genotype is the most important factor affecting somatic embryogenesis and organ...

Transgenic Robinia pseudoacacia plants were obtained by Agrobacterium tumefaciens mediated gene transfer. Agrobacterium strain LBA4404 harbouring a binary vector that contained the chimeric neomycin phosphotransferase II (NPTII) and beta-glucuronidase (GUS) genes was co-cultivated with hypocotyl segments of in vitro raised seedlings of Robinia. Parameters important for high efficiency regenerat...