Real-time PCR is a closed DNA amplification system that skillfully integrates biochemical, photoelectric and computer techniques. Fluorescence data acquired once per cycle provides rapid absolute quantification of ini...Real-time PCR is a closed DNA amplification system that skillfully integrates biochemical, photoelectric and computer techniques. Fluorescence data acquired once per cycle provides rapid absolute quantification of initial template copy numbers as PCR products are generated. This technique significantly simplifies and accelerates the process of producing reproducible quantification of nucleic acid molecules. It not only is a sensitive, accurate and rapid quantitative method, but it also provides an easier way to calculate the absolute starting copy number of nucleic acid molecules to be tested. Together with molecular bio-techniques, like microarray, real-time PCR will play a very important role in many aspects of molecular life science such as functional gene analysis and disease molecular diagnostics. This review introduces the detailed principles and application of the real-time PCR technique, describes a recently developed system for exact quantification of AUX/IAA genes In Arabidopsis, and discusses the problems with the real-time PCR process.展开更多
[ Objectives] The study was conducted to investigate the molecular identification of Salvia miltiorrhiza Bge. and its adulterants by DNA barcoding andspecific primer PCR. [ Methods] With ITS2 sequenceas DNA barcode, t...[ Objectives] The study was conducted to investigate the molecular identification of Salvia miltiorrhiza Bge. and its adulterants by DNA barcoding andspecific primer PCR. [ Methods] With ITS2 sequenceas DNA barcode, the materials were amplified by PCR and sequenced, and the NJ phylogenetic tree was constructed. The secondary structure of ITS2 was predicted by database and its website established by Koetschan et al. , and the self-designed primers were used to carry out specific primer PCR identification. [Results] ITS2 sequence length was around 470 bp. The results of cluster analysis showed that S. miltiorrhiza Bge. and its adulterants were clustered on different branches and showed monophyly. The comparison of secondary structure showed that S. miltiorrhiza Bge. had little differences from S. przewalskii, while there were significant differences from A. lappa in the number, size and location of stem-loop and the rotation angle of the spiral arm from the central ring. The specific primers could distinguish the S. miltiorrhiza Bge. and its counterfeits by PCR technique. [Conclusions] DNA barcoding and specific primer PCR are effective in distinguishing S. miltiorrhiza Bge. and its adulterants, and it has an important application foreground in the identification of Chinese herbal medicines.展开更多
文摘Real-time PCR is a closed DNA amplification system that skillfully integrates biochemical, photoelectric and computer techniques. Fluorescence data acquired once per cycle provides rapid absolute quantification of initial template copy numbers as PCR products are generated. This technique significantly simplifies and accelerates the process of producing reproducible quantification of nucleic acid molecules. It not only is a sensitive, accurate and rapid quantitative method, but it also provides an easier way to calculate the absolute starting copy number of nucleic acid molecules to be tested. Together with molecular bio-techniques, like microarray, real-time PCR will play a very important role in many aspects of molecular life science such as functional gene analysis and disease molecular diagnostics. This review introduces the detailed principles and application of the real-time PCR technique, describes a recently developed system for exact quantification of AUX/IAA genes In Arabidopsis, and discusses the problems with the real-time PCR process.
文摘[ Objectives] The study was conducted to investigate the molecular identification of Salvia miltiorrhiza Bge. and its adulterants by DNA barcoding andspecific primer PCR. [ Methods] With ITS2 sequenceas DNA barcode, the materials were amplified by PCR and sequenced, and the NJ phylogenetic tree was constructed. The secondary structure of ITS2 was predicted by database and its website established by Koetschan et al. , and the self-designed primers were used to carry out specific primer PCR identification. [Results] ITS2 sequence length was around 470 bp. The results of cluster analysis showed that S. miltiorrhiza Bge. and its adulterants were clustered on different branches and showed monophyly. The comparison of secondary structure showed that S. miltiorrhiza Bge. had little differences from S. przewalskii, while there were significant differences from A. lappa in the number, size and location of stem-loop and the rotation angle of the spiral arm from the central ring. The specific primers could distinguish the S. miltiorrhiza Bge. and its counterfeits by PCR technique. [Conclusions] DNA barcoding and specific primer PCR are effective in distinguishing S. miltiorrhiza Bge. and its adulterants, and it has an important application foreground in the identification of Chinese herbal medicines.
