BACKGROUND Multiple approaches for the site-directed mutagenesis (SDM) have been developed. However, only several of them are designed for simultaneous introduction of multiple nucleotide alterations, and these are time consuming. In addition, many of the existing multiple SDM methods have technical limitations associated with type and number of mutations that can be introduced, or are technica...

The ability of Shiga toxin-producing Escherichia coli (STEC) to cause severe illness in humans is determined by multiple host factors and bacterial characteristics, including Shiga toxin (Stx) subtype. Given the link between Stx2a subtype and disease severity, we sought to identify the stx subtypes present in whole genome sequences (WGS) of 444 isolates of STEC O157. Difficulties in assembling ...

A PCR assay for detection of enterovirus RNA in multiple specimen types from patients with neurological infections was evaluated. Combined PCR assay of cerebrospinal fluid and serum (systemic specimens) was more sensitive than assaying either specimen alone in children but not in adults. Compared with PCR in systemic specimens, detection of enterovirus RNA in throat swabs showed a sensitivity o...

Abstract Background The prevalence of gastrointestinal (GI) neoplastic polyps in Jack Russell terriers (JRTs) has increased Japan since the late 2000s. Recently, we demonstrated that JRTs with GI harbor identical germline variant APC gene (c.[462_463delinsTT]) heterozygous state. Thus, this disease is an autosomal dominant hereditary disorder. Although affected have distinct features, such as d...

Viruses are the leading cause of central nervous system (CNS) infections, ahead of bacteria, parasites, and fungal agents. A rapid and comprehensive virologic diagnostic testing method is needed to improve the therapeutic management of hospitalized pediatric or adult patients. In this study, we assessed the clinical performance of PCR amplification coupled with electrospray ionization-time of f...

The disease caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) in Wuhan, China, December 2019, has rapidly spread worldwide. SARS-CoV-2 is usually detected via real-time reverse-transcription polymerase chain reaction (RT-PCR). However, the increase specimen load institutions/hospitals necessitates a simpler detection system. Here, we present an ultra-rapid, RT-PCR assay for...

The potential for the non-coding intergenic rDNA spacer (IGS) to DNA fingerprint Giardia duodenalis isolates was investigated. Conserved PCR primers, specific for the flanking large and small rDNA genes, were used to amplify the IGS from 52 in vitro-cultured Giardia isolates. Four distinct IGS-PCR size groups (1.35-1.6 kb) were observed, which correlated closely with the major genetic assemblag...

is more sensitive but requires two steps [two rounds of amplification (PCR LDR) are needed]. The costs for setup are larger (PCR primers and three labeled oligonucleotides are needed per SNP) with respect to PCR/RFLP and allele-specific PCR but are similar for the TaqMan assay. The main advantage consists in the inherent multiplexing capability that can increase throughput, which decreases cost...

In this paper I tested whether Contradictory Matrix with 40 Inventive Principles, the simplest instrument from the Theory of Inventive Problem Solving (TRIZ), is a useful approach to a real-life PCR scenario. The PCR challenge consisted of standardization of fluorescence melting curve measurements in Competitive Amplification of Differentially Melting Amplicons (CADMA) PCR for multiple targets....

The article reported a novel methodology for real-time PCR analysis of nucleic acids, termed endonuclease restriction-mediated real-time polymerase chain reaction (ET-PCR). Just like PCR, ET-PCR only required one pair of primers. A short sequence, which was recognized by restriction enzyme BstUI, was attached to the 5' end of the forward (F) or reverse (R) PCR primer, and the new F or R primer ...