Skip to main content
Fig. 7 | Mobile DNA

Fig. 7

From: The SAMHD1-mediated block of LINE-1 retroelements is regulated by phosphorylation

Fig. 7

SAMHD1 does not impede ORF2p reverse transcriptase activity in LINE-1 RNPs. a The L1 RNP samples were purified from lysates of transfected 293T cells by ultracentrifugation through a sucrose cushion. The LEAP primer contains a linker sequence and was used to reverse transcribe L1 mRNA from RNPs. Reverse transcription occurred by either ORF2p (LEAP reaction) or exogenous MLV-RT. Synthesized cDNA was amplified by PCR with primers binding to the 3‘end of L1 cDNA and the linker sequence. b 293T cells were transfected with an empty vector (pcDNA) or the L1 expression vector pAD2TE1 together with expression plasmids for SAMHD1-myc T592A, SAMHD1-myc D207N, or ZAP-HA. Two days posttransfection, immunoblot membranes were probed with anti-T7 antibody (ORF1p-T7), anti-myc antibody (SAMHD1), and anti-HA antibody (ZAP). The amount of ORF1p in RNP samples after ultracentrifugation was determined by immunoblot using an anti-T7 antibody. c cDNA synthesized by either ORF2p or MLV-RT from L1 RNPs was analyzed by standard PCR with primers binding to the 3′ end of L1 and the linker sequence of the LEAP primer. Amplified PCR products were visualized on a 2% agarose gel

Back to article page