Interference No. 102,572
sequencing results . Holmes, ¶ 9 (CR-30). The fragment was purified by PAGE, cleaved
with Sau3A and the 182 bp fragment isolated by PAGE (fragment 2). Holmes, ¶ 9 .
Thereafter, fragments 1 and 2 were ligated together and the ligation reaction transformed
into E. coli. The resultant transformants were analyzed by restriction digestion and
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sequencing to confirm the construction of pKCEAInt1. Holmes, ¶ 10. To prepare
fragment 3, Holmes testified that he digested pK17G4 DNA with PstI and purified the
fragment by PAGE. This fragment was partially digested with AvaII, filled in and purified by
PAGE. This fragment was subsequently digested with HpaII and the 497 bp fragment
isolated by PAGE (fragment 3). Holmes, ¶ 11 (CR-31). For fragment 4, Holmes testified
that he digested the plasmid pKCEAInt1 with AvaI, filled in and digested with XbaI. The
large fragment was treated with BAP and isolated by PAGE (fragment 4). Holmes, ¶ 12.
The small fragment was digested with HpaII and the 169 bp fragment isolated by PAGE
(fragment 5). Holmes, ¶ 12 (CR-31). Fragments 3, 4 and 5 were ligated and the ligation
reaction transformed into E.coli. Resultant transformants were analyzed by restriction
digestion to confirm the construction of pKCEAInt2. Holmes, ¶ 13 (CR-31).
(10) Cabilly testified that he modified plasmid pKCEAtrp207-1* by cleaving out the
PstI-PvuI fragment from the ampicillin resistance gene, filling it in and relegating the blunt
ends to yield plasmid pKCEAtrp207-1*delta which is resistant to tetracycline but sensitive
12The Cabilly et al. brief (page 11, first paragraph) alleges that this analysis and
isolation of pKCEAInt1 was done on or about October 30, 1982. No one testified as to
this date. id.
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