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Figure 3 | BMC Biotechnology

Figure 3

From: Sequence specific visual detection of LAMP reactions by addition of cationic polymers

Figure 3

Precipitation titration of LAMP products by addition of PEI. (A) Effect of amount of PEI on sequence-specific incorporation of ROX-labeled lambda DNA recognition probes by DNA-PEI complex (Mw of PEI is 600). When 0.2 μmol to 1.0 μmol of PEI was added as a monomer, almost 100% of labeled probes hybridized to the LAMP products for lambda DNA was taken up by the DNA-PEI complex. On the other hand, when 0.4 μmol to 0.8 μmol of PEI was added as a monomer to a reaction solution in which an amplification reaction did not take place, a small amount (<20%) of labeled probes precipitated. Precipitation of this nonspecific probe did not take place when unrelated LAMP products (PSA) were present. (B) Effect of the Mw of PEI on sequence-specific incorporation of ROX-labeled lambda DNA recognition probes by DNA-PEI complex. When PEI with Mw 600 was used, the fluorescence intensity of supernatant decreased only when LAMP product for lambda DNA was present. As the Mw of the PEI used increased from 1,800 to 10,000, the fluorescence intensity of the supernatant decreased in the absence of a LAMP reaction since the formation of insoluble PEI-oligo DNA probe complex occurred. (C) Effect of amount of KCI on sequence-specific incorporation of ROX-labeled lambda DNA recognition probes by LAMP product-PEI (Mw = 600) complex. Normal LAMP reaction solution (control) contains 10 mM of KCI. Since formation of the PEI-LAMP product complex was inhibited by an increase in the amount of KCI added, the fluorescence intensity of supernatant increased regardless of the sequence of the LAMP product. (D) Effect of amount of LAMP product for lambda DNA on sequence-specific incorporation of ROX-labeled lambda DNA recognition probe by LAMP product-PEI complex (Mw of PEI = 600). Even when the amount of LAMP product was 1 μg per 25 μL, almost 100% of labeled probes (1 pmol) was taken up by PEI-DNA complex.

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