TaqMan® Technology and Real‐Time Polymerase Chain Reaction

Abstract

Twenty-eight PIC male piglets (similar birth and parity, weaned at 21±1.5 days) were used to study the effect of the different sources of dietary starch on the number of Bifidobacteria in the digesta of the duodenum, jejunum, ileum, cecum and colon. Pigs were randomly assigned to one of four diets formulated with corn starch, wheat starch, tapioca or pea starch. The determined ratio of amylose to amylopectin for these starches was 0.21, 0.24, 0.12 and 0.52, respectively based on the 16s rRNA sequences of maximum species of Bifidobacterium from GenBank to design the primers and probe. TaqMan polymerase chain reaction was developed to quantify the number of Bifidobacterium. We used this assay to detect genomic DNA of Bifidobacterium in the intestinal tract digesta of piglets, including duodenum, jejunum, ileum, cecum and colon. Our results indicated that, developed new real-time quantitative PCR assays can be allowed for rapid, convenient, reproducible and steady quantification of the Bifidobacterium in the intestinal content of piglets. Additionally, the present study revealed that high amylose/amylopectin ratio of starches significantly enhanced the numbers of Bifidobacterium in all segments of intestine.

International Collaborative Study of the Endogenous Reference Gene, Sucrose Phosphate Synthase (SPS), Used for Qualitative and Quantitative Analysis of Genetically Modified Rice

“…Quantitative Real-Time PCR Reactions. In this collaborative ring trial, we adopted the quantitative real-time PCR primers reported in our previous study and employed a new TaqMan probe with complementary sequence as our previous reported probe because there was a G at the 5 0 end of the previous probe (25 ), which could affect the signal intensity of fluorescence (dada not shown) (36 ). The sequences and locations of quantitative primers (SPS-1F/3R) together with the modified probe (SPS-P) are presented in Table 1 and Participants were requested to dilute each of these four DNA samples to the concentration of 10, 1, 0.1, 0.01, or 0.002 ng/μL by using the DNA solution provided.…”

Genetic diversity and linkage disequilibrium using SNP (KASP) and AFLP markers in a worldwide durum wheat (Triticum turgidum L. var durum) collection

“…The development of multiplexed array-based high-throughput genotyping technologies [21] or uniplex SNP genotyping platforms, such as TaqMan TM [22] and KASP TM , among others, has greatly facilitated the use of SNP markers. Kompetitive Allele Specific PCR or KASP TM is a cost-effective technology with high sensitivity, accuracy and flexibility allowing genotyping either few samples with many SNP markers or several samples with few SNP markers in a single plate [11].…”

Clinically relevant cancer biomarkers and pharmacogenetic assays

“…Alternatively, RT-PCR is based upon the generation of a fluorescent signal during the PCR process, which is detected during cycling and reflects the amount of PCR product synthesized during the reaction. 33 RT-PCR has better sensitivity than Sanger sequencing but only identifies mutations that the sequence specific primer is designed to detect. 32 PCR-based methods are also utilized to detect candidate genes as a research tool in validating genes discovered through genome-wide association studies.…”