Homogeneous and one-step fluorescent allele-specific PCR for SNP genotyping assays using conjugated polyelectrolytes

In vitroselection of DNA-cleaving deoxyribozyme with site-specific thymidine excision activity

“…Many of these so-called single-nucleotide polymorphisms (SNPs) have been linked to human diseases, including phenylketonuria, hemophilia and certain cancers (6). Significant advances have been made in the past several years to develop accurate, rapid and cost-effective technologies for SNP detection, such as denaturing gradient gel electrophoresis (7), microfluidic devices (8), technologies based upon chip (9), allele-specific polymerase chain reaction (PCR) (10), strand displacement amplification (11), rolling circle amplification (12) and ligase chain reaction (13). …”

RNA-primed allele-specific PCR

“…Clones were picked from plates (100 mg mL 1 ampicillin) and grown in 40 ml LB medium (100 mg mL 1 ampicillin) until the OD600 was about 1.0. TaqM1 polymerase was expressed by inducing with 2 mM IPTG for 12 h. After centrifugation and washing three times with 1 × Taq buffer [20 mM Tris-HCl (pH 8.4), 20 mM KCl, 10 mM (NH 4 ) 2 SO 4 , and 2 mM MgSO 4 ], lysis was achieved using a 500 mL 1 × Taq buffer containing 1 mg mL 1 lysozyme. The lysate was incubated at room temperature for 30 min followed by heat denaturation of all nonthermostable proteins at 75 °C for 15 min.…”

“…The increasing need for large-scale genotyping applications of single-nucleotide polymorphisms requires the development of low-cost technologies accessible to minimally equipped laboratories. Allele-specific polymerase chain reaction (AS-PCR) is a convenient and inexpensive method for genotyping single-nucleotide polymorphisms (SNPs) and mutations [3][4][5]. Based on the SNP in the genome, the AS-PCR primers were designed with specific mismatches at the 3-end that allowed preferential amplification of one allele relative to another on account of the primers being complementary to the SNP site.…”

DNAzyme based gap-LCR detection of single-nucleotide polymorphism

“…SNPs are also thought to be key enablers in realizing the concept of personalized medicine. For this reason, over the past several years a great deal of effort has been devoted to developing accurate, rapid, and cost-effective technologies for SNP analysis from restriction endonuclease mapping approach (Wilson et al, 1982), denaturing gradient gel electrophoresis (Borresen-Dale et al, 2001;Myers et al, 1985), RNase or chemical cleavage of mismatched heteroduplexes (Cotton and Grompe, 2001;Cotton et al, 1988;Theophilus et al, 1989) to newly developed techniques such as allele-specific PCR (Cook et al, 2012;Duan et al, 2009;Liu et al, 2012;Muneta et al, 2012;Myakishev et al, 2001), strand displacement amplification (SDA) (Shi et al, 2011;Van Ness et al, 2003;Wang et al, 2011Wang et al, , 2012aWang et al, , 2012b, rolling circle amplification (RCA) (Bi et al, 2010;Li et al, 2010;Wang et al, 2011Wang et al, , 2012aWang et al, , 2012bWu et al, 2010;Zhang et al, 2009), and technologies based upon chips (Tanaka et al, 2012;Van Bers et al, 2012) and microfluidic devices (Hatakeyama et al, 2009;Ng et al, 2007;Russom et al, 2003). However most of those techniques incorporate primer extensions or PCR assays to obtain adequate amounts of starting material.…”