Identification of Armeniacae Semen Amarum Adulterated in Persicae Semen by Allele-specific Polymerase Chain Reaction / 中国实验方剂学杂志

Objective:Due to the limitation of traditional identification methods of Chinese medicinal materials， the study established a rapid method to identify Persicae Semen mixed with Armeniacae Semen Amarum by allele-specific polymerase chain reaction （PCR）. Method:By comparing the ribosomal DNA internal transcribed spacer （ITS） gene sequences of Persicae Semen and Armeniacae Semen Amarum， single nucleotide polymorphism （SNP） sites were searched and specific primers were designed. Different Persicae Semen and Armeniacae Semen Amarum samples were amplified by PCR， the effects of annealing temperature， primer concentration and cycle number on the PCR reaction system were optimized， and the specificity and detection limit of this method were investigated. In addition， the established PCR method was used to detect the samples of Persicae Semen mixed with different proportion of Armeniacae Semen Amarum from different sources and producing areas. Result:A specific PCR method for identifying Persicae Semen mixed with Armeniacae Semen Amarum was established. When the annealing temperature was 63 ℃ and the number of primer cycles was 30， only Armeniacae Semen Amarum could be amplified with 432 bp specific band， while Persicae Semen samples did not have this band. The minimum detection limit of this method for Armeniacae Semen Amarum was 0.2 ng， and the detection limit for Armeniacae Semen Amarum adulterated in Persicae Semen was 1%. Conclusion:The established allele-specific PCR method can accurately detect whether there is Armeniacae Semen Amarum in Persicae Semen， which can provide experimental basis for the quality control of Persicae Semen and guarantee the safety of its clinical use.

Identification of Levisticum officinale Adulterated in Angelica sinensis by PCR-RFLP / 中国实验方剂学杂志

Objective:To establish a rapid method to identify <italic>Levisticum officinale </italic>adulterated in<italic> Angelica sinensis</italic> by polymerase chain reaction -restriction fragment length polymophism（PCR-RFLP）. Method:By comparing sequences restriction sites in ribosomal DNA Internal Transcribed Spacer（ITS） of <italic>A. sinensis</italic> and <italic>L. officinale</italic>，the specific restriction site Fnu4HI of <italic>L. officinale</italic> was selected，and the primers for PCR-RFLP reaction were designed. Different <italic>A. sinensis</italic> and <italic>L. officinale</italic> were amplified by PCR. The conditions affecting the PCR-RFLP reaction，such as annealing temperature，primer concentration，cycle number and enzyme digestion reaction time，were optimized，and the accuracy of the method was investigated. The established PCR-RFLP identification method was used to investigate the applicability of <italic>L. officinale </italic>adulterated<italic> in A. sinensis</italic> with different aduleration ratios and different origins. Result:A PCR-RFLP method for identifying <italic>A. sinensis</italic> mixed with <italic>L. officinale</italic> was established. When the annealing temperature was 62 ℃ and the number of cycles was 30，when the <italic>L. officinale </italic>adulterated in<italic> A. sinensis</italic> could be digested by Fnu4H I restriction endonuclease after amplification with specific primers，and the two single DNA bands were detected between 100-500 bp，the <italic>A. sinensis</italic> were all negative. The minimum detection limit of this method for adulterated <italic>L. officinale</italic> in <italic>A. sinensis</italic> was 3%，which could be used for the detection of adulterated <italic>L. officinale</italic> in <italic>A. sinensis</italic>. Conclusion:The established PCR-RFLP identification method is sensitive and accurate in detecting whether there is <italic>L. officinale</italic> in <italic>A. sinensis</italic>，and it provides inspection reference and basis for the quality control of <italic>A. sinensis</italic>，with great significance to ensure the safety of its clinical medication.