Quantification of Mouse Genome in Xenografts of Human Pancreatic Cancers by Species-Specific Length Polymorphisms
MT Lin, LH Tseng, H Kamiyama, M Kamiyama, P Lim, JR Eshleman. Johns Hopkins University School of Medicine, Johns Hopkins Hospital, Baltimore, MD; National Taiwan University Hospital, Taipei, Taiwan
Background: Establishing cancer cell lines from primary tissues is essential for many research areas of cancer biology. Isolation of human cancer cell lines, from pancreatic cancers and other solid tumors, is surprisingly difficult, primarily due to overgrowth of fibroblasts and other mesenchymal stromal cells during the early stages of in vitro culture. Cancers can be maintained by serial xenografting in athymic mice or severe combined immunodeficient (SCID) mice.
Design: We have developed a molecular detection assay to monitor the human/mouse genomic chimerism from DNA samples extracted directly from xenografts and from human pancreatic cell lines established in vitro after explanting the tumors. The assay was designed based on deletion/insertion variation between human and mouse genomes, a strategy similar to the use of amelogenin gene polymorphism to determine human female/male chimerism. The percentage of mouse DNA was calculated according to the relative peak height of PCR products analyzed by capillary electrophoresis.
Results: Three markers from chromosomes 9 and 10 showed accurate prediction of mouse genome ratio and were successfully combined into a multiplex PCR reaction. The assay was used to quantify 24 mouse xenografts of human pancreatic cancers. The results of chimerism analysis were also used to select flasks with highest percentage of human cells for subsequent passage of cultures and to confirm the purity of pancreatic cancer cell lines for subsequent biomedical studies.
Conclusions: In the current study, we have developed a PCR assay for rapid and robust quantification of mouse genome in xenografts of human pancreatic cancers. The results facilitate DNA sequencing of human pancreatic cancers and in vitro establishment of human pancreatic cancer cell lines.
Category: Liver & Pancreas
Wednesday, March 11, 2009 9:30 AM
Poster Session V # 244, Wednesday Morning