Additional file 1 Evaluation of the number of integrated copies. A. 3.5 kb. AgeI. P-HTG. 2.0 kb. 1.6 kb. ATG. HTG. ∆neo. 1.4 kb. 0.9 kb. P SV40. P SV40. P SV40. P SV40. wt 1* 2* 3 4* 5. B. 21 kb. AgeI. 5.0 kb. 2.0 kb. 0.9 kb.
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Evaluation of the number of integrated copies
wt 1* 2* 3 4* 5
- wt 1 2 3* 4 5* 6 7 8* 9 10 11 12
A: Analysis of bordering fragmentsfrom P-HTG tagged HEK293 cells.
lanes 1-5 individual clonesobtained upon tagging.
B: Analysis of bordering fragmentsof P-Ab-HTG antibodytagged CHO cells.
lanewt, untransfected CHO-K1 cells, lanes 1-12 correspondtoclones #1-12 in Fig. 2B.
Cloneswithsinglebandsaremarkedwith an asterisk.
wt: untransfectedcells; theneo probe usedforhybridizationisindicated.
Foranalysis, highmolecular DNA of individual clones was extractedaccordingto Ramirez-Solis et al., 1992. The DNA was cutwithAgeIandsubjectedto Southern Blotting. Fordetectionof bordering fragments, theblot was hybridizedtotheNeosequence. Each band correspondsto an individual integrationsite. Single bandsindicatesingleintegrationsites.
Ramirez-Solis, R., et al. (1992). Genomic DNA microextraction: a method to screen numerous samples. Anal. Biochem. 201: 331–335.