Strand-specific Southern blot was performed as described6 (link). Briefly, AflIII and AseI digested samples were separated on a 7% polyacrylamide gel and transferred to a nylone membrane (Hybond-N+, Amersham). After transfer, the membrane was rinsed in 4X SSC for 5 minutes, and UV irradiated to crosslink the DNA to the membrane. The membrane was then pre-hybridized with 25 ml hybridization buffer (ULTRAhyb from Ambion) for at least 3 hours at 42 ° C. Strand-specific probes generated by a PCR based primer extension reaction6 (link), were added to the hybridization buffer and incubated with the membrane at 42 ° C overnight. After overnight hybridization, the membrane was washed 2 times with 2X SSC, 0.1% SDS for 5 minutes at 42 ° C. The dried membrane was exposed to a phosphorimager screen.
Uncropped images of gels and autoradiographs used in this study can be found inSupplementary Data Set 1 .
Uncropped images of gels and autoradiographs used in this study can be found in
