Cloning and expression of soluble truncated variants of Borrelia

Details Cloning and expression of soluble truncated variants of Borrelia Cloning and expression of soluble truncated variants of Borrelia

1992-09-08

1996-11-05

Jones, W. Gary

Chemistry: molecular biology and microbiology

Micro-organism, per se ; compositions thereof; proces of...

Bacteria or actinomycetales; media therefor

4353201, 530359, 530402, 536 237, C12N 120, C12N 1570, C07K 1300, C07H 2104

Patent

active

055717187

ABSTRACT:
A method is provided herein for preparing soluble recombinant variations of Borrelia lipoproteins such as Borrelia burgdorferi outer surface protein A (OspA) and outer surface protein B (OspB), and B. hermsii variable major protein 7 (Vmp7). The method includes synthesizing a set of oligonucleotide primers, amplifying the template DNA utilizing the PCR, purifying the amplification products, cloning the amplification products into a suitable expression vector, transforming a suitable host utilizing the cloned expression vector, cultivating the transformed host for protein production and subsequently isolating and purifying the resulting protein. Also provided are soluble, recombinant variations of Borrelia burgdorferi outer surface protein A (OspA), outer surface protein B (OspB), and B. hermsii variable major protein 7 (Vmp7). The expression vectors harboring DNA encoding the recombinant variations, pET9-OspA, pET9-OspB and pET9-Vmp7, as well as the E. coli host BL21(DE3)/pLysS transformed with each of these vectors, are also disclosed.

REFERENCES:
Sam Brook et al. Molecular Cloning: A Laboratory Manual, 2nd ed. Cold Spring Harbor Laboratory Press, N.Y. (1989) pp. 8.46-8.49.
Wu, in Bacterial Outer Membranes as Model Systems, ed. Inouye John Wiley & Sons, New York, 1987, pp. 37-50.
Brandt et al., Infection & Immunity, vol. 58(4)*, pp. 983-1991 (Apr. 1990).
Howe et al. (8 Feb. 1985) Science, vol. 227, pp. 645-646.
Lin et al. (1980) Journal of Biol. Chem. vol. 255(3), pp. 1160-1163.
Kitten et al ., Proc. Natl. Acad. Sci., vol. 87, pp. 6077-6081, Aug. 1990.
Fikrig, et al., "Protection of Mice Against the Lyme Disease Agent by Immunizing with Recombinant OspA", Reports, Oct. 26, 1990.

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