Appeal No. 1999-1425
Application No. 08/393,321
b) separating the disrupted bacterial cell debris from the liquid containing intact
plasmid DNA.
10. A mechanical method for disruption of plasmid-containing bacterial cells and
release of intact plasmid DNA, comprising the steps of:
a) passing liquid containing plasmid-containing bacterial cells through a bead mill
containing beads of about 0.1 mm to about 1 mm in diameter, at an agitation speed of
about 1,000 to 2,500 rpm, wherein the liquid is processed in the bead mill, for a total
residence time in the bead mill of at least about 3 minutes, whereby bacterial cells are
disrupted and intact plasmid DNA is released; and
b) separating the disrupted bacterial cell debris from the liquid containing intact
plasmid DNA.
The references relied upon by the examiner are:
Agerkvist et al. (Agerkvist) “Characterization of E. Coli. Cell Disintegrates from a
Bead Mill and High Pressure Homogenizers,” Biotechnology and Bioengineering,
Vol. 36, pp. 1083-1089 (1990)
Sambrook et al.(Sambrook) “Extraction and Purification of Plasmid DNA,”
Molecular Cloning: A Laboratory Manual, 2nd Edition, Cold Spring Harbor Press
Sect 1.21-1.52 (1989)
Sauer et al. (Sauer) Disruption of Native and Recombination Escherichia Coli in a
High-Pressure Homogenizer,” Biotechnology and Bioengineering, Vol. 33,
pp. 1330-1342 (1989)
Johnson “Isolation and Purification of Nucleic Acids,” Nucleic Acid Techniques in
Bacterial Systematics, Stackebrandt et al., eds., John Wiley & Sons, New York, NY., pp. 1-
19 (1991)
Grounds of Rejection
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