Figure 5.

hpol η, instead of other TLS polymerases, plays a key role in reverse transcription in human cell extracts. Extracts of CRISPR-Cas9 knock-out cell lines were made, and each cell extract (1.4 mg protein/ml) was incubated with 1 μm DNA/DNA (dG), DNA/DNA(CPD), or DNA/RNA (rG), respectively, in the presence of a dNTP mixture (the concentration of each dNTP was 450 μm). An unrelated annealed dsDNA-RNA hybrid and ssDNA (100 μm concentrations) were used to prevent degradation. The reactions were conducted for 10, 30, and 60 min. See “Oligonucleotide substrates” under “Experimental procedures” for the oligonucleotide sequences used.