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Principle - ACP Structure and Rationale
Numerous approaches have been introduced to increase primer annealing
specificity. Longer primers with universal, homopolymer, or loop
sequence tails at their 5' ends improve the specificity of PCR amplification
and the stability of hybridisation. However, these approaches do
not abrogate nonspecific hybridisation. resulting from the involvement
of the tail or loop sequence in the priming reaction. However, the
ACP specifically targets sequences hybridisation. to the template
via a regulator linker. This regulator generates a region with a
lower melting temperature via the formation of a bubble-link structure
at specific temperatures. Essentially the ACP linker prevents the
nontarget universal sequence from annealing under conditions in
which the target 3' sequence anneals to the template.
1) ACP Structure |
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Fig. 1. ACP™ Structure. The
ACP comprises: 1. a 3'-end core portion (targeting) having
a hybridising sequence substantially complementary to a site
on a template nucleic acid to hybridise therewith; 2. a 5'-end
portion having a universal sequence; and 3. a regulator bridging
the core and universal sequences of ACP which plays a key
role in annealing of each portion to a template. |
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Fig. 2. Rationale of ACP Technology |
2) Rationale
| ACP
- The regulator specifies that only the target core sequence
of ACP can hybridise to the template.
- The non-target universal sequence of ACP prevents the
hybridisation. to the template by the regulator as a bubble-like
form.
- Therefore, the specific hybridising strength of the target
core sequence can be maximised, resulting in only target
specific products.
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Conventional long primer
- Conventional longer primer cannot restrict the hybridisation.
sequence.
- Therefore, the non-target universal sequence is involved
in the hybridisation. to the template.
- The conventional longer primer can bind to non-target
sequences of the template due to the involvement of the
non-target universal sequence in the hybridisation., resulting
in non-target products.
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