Definition

Annealing Control Primer (ACP)™ Technology (patent pending) represents the most accurate and extensive PCR technology developed by Seegene Inc.

ACP™ Technology, a novel method which completely eliminates false-positive results, uses specially designed primers that anneal specifically to the template and allows only genuine products to be amplified. The ACP system is based on two principles; the unique tripartite structure of the primers, which have the 3’ end target core sequence and the 5’end nontarget universal sequence which are separated by a regulator linker, and the interaction of each region during two-stage PCR.

Applications

• Amplification of a target nucleic acid sequence
• Differentially expressed gene (DEG) discovery
• Diagnosis / Genomic fingerprinting for polymorphism
• SNP Genotyping / Multiplex PCR
• Full-length cDNAs / 5'-RACE and 3'-RACE

Features

Primer design is the most critical factor in the current PCR methods to have a primer bind to a target specific site.

The specificity with which a primer anneals only to its target (and not non-target) sequence is the most critical factor in determining the success of PCR amplification. ACP™ Technology provides a primer with annealing specificity to the template and allows only real products to be amplified, such that it enables the researchers to find only real products as a result. The principle of ACP™ Technology is based on the unique tripartite structure of a particular oligonucleotide primer having 3'- and 5'-end distinct portions separated by a regulator and the interaction of each portion during two stage PCR amplification.

• NO Non-target! Only Target sequence!

  The regulator bridging the 3'- and 5'-end portions restricts a primer annealing portion to the 3'-end core portion during the 1st stage PCR, so that the annealing sequence of a primer can be precisely controlled, which make it possible to design a primer with a desired number of annealing sequence. This is particularly useful when an annealing portion of a primer has to be limited (e.g., single nucleotide polymorphism (SNP) genotyping, DNA microarray screening, and detection of differentially expressed genes).

• NO Non-specific Product!

  The regulator bridging of the 3'- and 5'-end portions interrupts the annealing of the 5-end portion to the template, and eventually the 5'-end portion not involved in the annealing provides the 3'-end portion with primer annealing specificity.

• Detect One Base Mismatching!

  The specificity of primer annealing is sensitive enough to detect even a single base mismatching. It is particularly useful for the identification of a nucleotide variation in a target nucleic acid, including, single nucleotide polymorphisms and point mutations.

• NO Non-specific Product through 2 step!

  Non-specific annealing is excluded from the amplification of the 1st primed PCR product by using only the sequences at the 3'- and 5'-ends of the 1st PCR product as donors of priming sequences.

• Great target product!

  The ACP™ system utilises a two stage PCR (ACP™ based PCR) which allows the products to be excluded from non-specific amplifications.

  Ordinary, high specificity in PCR is favoured by following:

  • Optimal concentration of Mg2+, other ions, primers, dNTPs and Taq DNA polymerase
  • Choice of thermostable DNA polymerases
  • Primer design
  • Efficient denaturation, high annealing temperatures and fast ramping rates
  • Limiting the number of cycles and their length
  • Thermal cycler efficiency
  • PCR additives such as DMSO, betain, single-stranded DNA binding proteins, enhancer (Stratagene), Q-solution (Qiagen), etc.
  • Template quality
  • Touchdown PCR, hot start PCR, booster PCR and nested PCR.

ACPs tolerate the above parameters. Thus, ACP-based PCR generates high specificity regardless of the above parameters.