A rapid RT-PCR based method to isolate complementary DNA fragments flanking retrovirus integration sites.
Proto-oncogenes in retrovirally induced myeloid mouse leukemias are frequently activated following retroviral insertion. The identification of common virus integration sites (VISs) and isolation of the transforming oncogene is laborious and time consuming. We established a rapid and simple PCR based procedure which facilitates the identification of VISs and novel proto-oncogenes. Complementary DNA fragments adjacent to retrovirus integration sites were selectively isolated by applying a reverse transcriptase (RT) reaction using an oligo(dT)-adaptor primer, followed by PCR using the adaptor sequence and a retrovirus long terminal repeat (LTR) specific primer. Multiple chimeric cDNA fragments suitable for Southern and northern blot analysis were isolated.
Valk
P J
PJ
Institute of Hematology, Erasmus University Rotterdam, The Netherlands.
Joosten
M
M
Vankan
Y
Y
Löwenberg
B
B
Delwel
R
R
D000818:Animals; D017931:DNA Primers; D018076:DNA, Complementary / Q000032:analysis* / Q000235:genetics; D007951:Leukemia, Myeloid; D051379:Mice; D016133:Polymerase Chain Reaction / Q000379:methods*; D011519:Proto-Oncogenes / Q000235:genetics; D012190:Retroviridae / Q000235:genetics*; D014407:Tumor Cells, Cultured; D016662:Virus Integration / Q000235:genetics*
D017931:DNA Primers; D018076:DNA, Complementary