Figure 9. Cleavage of a radioactively
labeled single- and double-stranded DNA by tear lipocalin and EndoA. A:
A 32-mer oligonucleotide was digested by TL (lanes 1–3), the minor tear
endonuclease (lanes 5–7), or DNase I (lanes 6–9) at 5, 10, and 20 min,
respectively. The lane designated M shows oligonucleotide sizing
markers. The size of the single strand digestion products differ for
the three enzymes. B: A 211 bp PCR product was digested by TL
(lanes 1, 2, 3), DNase I (lanes 1’, 2’, 3’), or the minor endonuclease
(lanes 1”, 2”, 3”) in 10, 20, and 30 min, respectively. The lane
headings, A, T, G, and C, identify the respective base-specific
sequencing reactions. Sequencing was performed using the dideoxy method
with T7 DNA-polymerase. The corresponding sequence of the dsDNA is
shown (left). The varying intensity of the bands at the sequences shown
reflect different rates of cleavage related to relative sequence
preferences for each of the three enzymes.