Appeal 2007-0082
Application 10/171,498
for "regenerating ordinary corn protoplasts into fertile plants". (Science, p. 630,
col. 1 and 2). The Science article indicates the first problem was solved by the use
of a "gene gun … to propel genes directly into whole cells." (Science, p. 630, col.
2). The second problem was also solved because the gene gun procedure resulted
in cells that were "amenable to cell culture procedures." (Science, p. 630, col 3).
When these two problems were solved, researchers obtained fertile genetically
engineered corn plants expressing a gene for herbicide resistance. (Science, p. 630,
col. 2).
As discussed above, these are the very obstacles addressed at length in
Tomes.
Again, Tomes states that,
[a] newly developed method of introducing genetic material into plant
cells is particle gun technology, also called microprojectile or
microparticle bombardment, which involves the use of high velocity
accelerated particles… It has been demonstrated that such
microparticles can enter plant cells without causing death of the cells
and that they effectively deliver foreign genetic material into intact
epidermal tissue of certain plants.
(Tomes, col. 3, ll. 14-30). Tomes further states that the
time within the subculture interval is an important determinant of the
efficiency of stable transformation. The preferred time for
microparticle bombardment is one day after subculture. … The callus
is prepared for bombardment by passing through a 700 micron sieve
which breaks the larger clumps which have a maximum diameter of
about 650 microns. … Preferably the cells are bombarded on the first
or second day following sieving.
It was also determined that both sieved callus and suspension cultures
require a certain density on the Petri dish and a specific surface area for
treatment with bombardment if successful transformation is to be achieved.
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