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Origin of tissue from which ~ wa, isolated. Rder~c~rcino,~o, C; M, (~entlsl normal colonic mJcoso; P, polyp; LM, r~tosto$i$ to lung; ! u~h~cy re. Total number of G418r~ colonies plated in ~elective ~edia. I~, r~t detor~ined. Scruples were scored l~i tiv@ fop fccu,J-for~<~tio~ i f )6 f~ilcel I ~puloti~ ~i~ int~iote ~s ~e listed ~ +/-. S~i~ were ol~ s~ f~ tM ~e of collies of ~olif~oti~ cells in t~ ~lete ~i~, ~ in ~di~ fr~ ~i~ ~, E~ ~ ~n ~itt~. ~le ~e 5 Cell line~ derived fros pri~:r~ tronsf$ctions that ~eee used to isolate Ot4R for r~c~'K~-y trc~fection oxp~rim~nt~. • ~is s~lectio~ w~s also ~ in tM ¢~b~ of in~mJl~n. 50188988

?i~as, Ja=es .~i. Table 2 Secondary Transfecfion Assays Selection1 G418r Colonies Tested focus. Comvlete -PDGF Selected -EGF cell lines PDI (-P1) 1055 ND ND + W'I'S c (-P1) 500 ND ND +/- PDm (-IE1) ND + ND ND llCA 490 +/- +/- 11CB 650 - - + 13EA 1000 - ND ND 1SPA 490 + +/- + ND 40 ND 41 +/- 42 N'D 43 26 50188989

~-alls / dish 50188970

Legend to Figure 1 Cells were seeded at 1 x 10~ cells/3s mm dish and maintained in the complete serum-free medium (black bars), or in medium lacking PDGF (stipled bars). Cell numbers were determined 9 days (A) or 14 days (B) after.plating. 27 50188971

Pipas, J~es ~I. 29. 50188972

Pip~s, Legend to Figure 2 Cell l~ne P48 is a secondary transfectant derived from a focus induced by DNA isolated from the a~pparently normal mucosa of a patient with Gardner syndrome. DNA from the P48 line was d~gested with EcoRl, BamHl, ¢~r Hind/I/ and electrcphoresed through a 1% agarose gel, transferred to nitrocellulose and hybridized against a human H~'as specific probe. The two lanes on the right show hybridization to a BamH1 digest of normal human DNA and to NIH3T3 cell DNA. 3O 50188973

Pipas, Ja~es ~. ~ 1.3 kb 2 31 50188974

LEGEND TO FIGURE 3 Lymphocyte DNA isolated from a patient with Gardner syndrome was used to transfect NIH3T3 cetls. DhlA Lsolated from colonies grown under selective conditions was used to obtain secondary transformants. 20 p.g of DNA from two secondary transformants was digested with EcoR/, resolved on ~ 0.8% agarose gel, and transferred to nitrocellulose. The filter was hybridized to & human a/u-specific probe labeled and by nick translation to a specific activity of 2.2 x 1G3 dprrdlJ.g. The filter was washed in conditions corresponding to Tm-27°C., and exposed for 20 hours on XAR-5 film with a Lighting Plus intensifying screen. 3-" 50188975

1 2 :33 50188976

Pipaa, Ja~.s ~. Legend to Figure 4 Southern hybrldizat~on of high molecular weight DNA from serum-free selected ceil lines to the human Alul repetitive sequence. DNA was digested with EcoRI, • eiectrophoresed on a 1% agarose gel, transferred to nitrocellulose and hybridaized to the human.specific sequences present in the BLUR8 probe. Lane 1 and 2 contain DNA from cell line 40 and P86 respectively. 50188977