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10 In Situ Hybridization for RNA: Nonradioactive Probe: ss cDNA Probe
149
(a)
singlestranded
+
10
5
doublestranded
+
(b)
singlestranded
+
Fig. 2. Analysis of the final products of two-step (a) and single-step (b) asymmetric peR on hydroxyapatite column. The fractions of the major peaks were
evaluated by electrophoresis and filter hybridization. dsDNA (first peak) and
ssDNA (second peak) were eluted separately.
(a)
o
(b)
--- ____ 5-._--.
____ product (ssDNA)
-template (dsDNA)
10
20
30
40
50 55 0 cycles
x1
x 1/10
x 1/100
Fig. 3. Electrophoresis and filter hybridization of the second peR product. 55
cycles of the peR were performed and the product was sampled every 5 cycles.
The samples were electrophoresed (a), or applied to the membrane and hybridized with the complementary oligonucleotide probe (b). The peR products
increased over 50 cycles.