ABSTRACT
This study aimed to identify Inulae Herba, Inulae Flos and their closely related species using the ITS2 bar-code, and secure the quality and clinical curative effect of these medicinal materials. DNA was extracted from Inula linariifolia, I. japonica, I. britanica, which are the original species of Inulae Herba and Inulae Flos, together with their closely related species. The ITS2 sequence was amplified by PCR and sequenced bidirectionally. Sequence assembly and generation of consensus sequence were conducted by the CodonCode Aligner. The genetic distances were comput-ed using MEGA 5.0 in accordance with the Kimura-2-parameter (K2P) model, and the phylogenetic tree constructed by the neighbor-joining (NJ) method. The results showed that the ITS2 sequences of the various species have stable variable sites. The intraspecific genetic distances among Inulae Herba and Inulae Flos were obviously lower than the interspecific genetic distance among the above two medicinal materials and its adulterants. The NJ tree based on ITS2 sequences can clearly differ from Inulae Herba, Inulae Flos and their closely related species. It is concluded that ITS2 sequence can be used as DNA barcode to identify Inulae Herba, Inulae Flos and their closely related species.
ABSTRACT
This study aimed to design a pair of primers for amplifying internal transcribed spacer 2 region which was used to identify Gastrodia elata through optimizing of DNA extraction and PCR amplification process. Two sequences are used for research object by amplification of universal primers. Design three pairs of specific primers by Primer Premier 5.0 and select the highest specificity through the study of 22 samples. The results showed that identification efficiency of the primer named TM2F-2R is as high as 90.9% when Annealing Temperature is equal to 54 degrees Celsius. Therefore, TM2F-2R can be used as primers ITS2 sequences of G. elata, this article provides a set of accu-rate and stable identification methods for G. elata in the molecular identification.
ABSTRACT
In this study, the internal transcribed spacer 2 of nuclear ribosomal DNA (ITS2) sequence was used for i-dentifying A nemone raddeana and its adulterants to ensure the quality of medicines and clinical efficacy. Genomic DNA was extracted from 36 samples using Genomic DNA kit and used as templates for Polymerase Chain Reaction (PCR). Sequence assembly and consensus sequence generation were performed by CodonCode Aligner. The intraspe-cific and interspecific genetic distances were computed and the neighbor-joining tree was constructed by MEGA 5.1 in accordance with the Kimura 2-Parameter (K-2P) model. Results: The length of ITS2 sequence of A nemone rad-deana was 216 bp. The Maximum intraspecific genetic distance was 0.014, the minimum interspecific genetic dis-tance was 0.021. The NJ tree showed that A . raddeana differ from its adulterants obviously. Conclusion: ITS2 se-quence was able to identify A . raddeana and its adulterants correctly stably and correctly, which provides a new tech-nique to its identification.
ABSTRACT
Objective: To identify Xanthii Fructus and secure its quality and safety in medication. Methods: Total ge-nomic DNA was extracted from Xanthii Fructus and its adulterants. ITS2 sequences were amplified, and purified PCR products were sequenced. Sequence assembly and consensus sequence generation were performed using the CodonCode Aligner V 4.2. The Kimura 2-Parameter (K2P) distances were calculated using MEGA 5.0. The neigh-bor-joining (NJ) phylogenetic trees were constructed. Results: The intraspecific genetic distances of Xanthii Fructus were 0. The interspecific genetic distances between Xanthii Fructus and its adulterants were ranged from 0.009 to 0.542. The NJ tree showed that Xanthii Fructus could differ from its adulterants obviously. Conclusion: ITS2 can be used to identify Xanthii Fructus from its adulterants effectively, and our study further confirmed the effectiveness of ITS2 to identify traditional Chinese medicinal materials.
ABSTRACT
Psammosilene Radix is a famous miao national herb and it is rare and endangered nowadays. However, it is often confused with the root of Silene viscidula. In this study, the ITS2 regions was used to identify Psam-mosilene Radix and its adulterants. All DNA samples of Psammosilene Radix and its adulterants were extracted. The ITS2 regions were amplified and sequenced, and the final sequences were assembled using the CondonCode Aligner. The genetic distances were calculated by kimura 2-parameter (K2P) model and the Neighbor-Joining(NJ) phylogenetic trees were constructed using MEGA5.1.BLAST 1, the nearest distance and phylogenetic tree (NJ-tree)methods were used to assess the identification efficiency of the ITS2 region. Results indicated that the length of ITS2 sequences of Psammosilene Radix w 229 bp, the identification effficiency of ITS2 region using BLAST 1 was 100%;and the maximum intra-specific K2P distance were lower than the minimum inter-specific K2P distance. Additionally, the NJ tree based on ITS2 sequence indicated that Psammosilene Radix and its adul-terants could be distinguished clearly . In conclusion , the ITS2 region as DNA barcodes could identify Psammosi-lene Radix and its adulterants stably and accurately. Furthermore, the application of ITS2 barcode in the identifi-cation of TCM has a good prospective .
ABSTRACT
In this study,Polygalae radix and its adulterants were identified by psbA-trnH sequence.The genomic DNA was extracted from forty-six samples, the psbA-trnH sequences were amplified and sequenced Bi-directionally, and then assembled sequences by Codoncode Aligner V 3.7.1. The genetic distances were computed by kimura 2-parameter (K2P) model, and the Neighbor-Joining tree was constructed by MEGA 6.0. Results showed that minimum intra-specific K2P distance of Polygala tenuifolia and Polygala sibirica were 0.004 and 0, which were smaller than the maximum intra-specific K2P. The NJ tree showed Polygalae radix can be distinguished from its adulterants by psbA-trnH sequences. Therefore, using psbA-trnH sequences can distinguish Polygalae radix from its adulterants.
ABSTRACT
In this study, the ITS2 sequence was used to identify Pinelliae Rhizoma and its adulterants to ensure its market circulation, clinical effect and safety. All genomic DNA from 59 samples were extracted successfully. The Kimura 2-Parameter (K2P) distances and NJ tree were calculated using software MEGA 6.0. The length of the ITS2 sequence of Pinelliae Rhizoma was 251 bp. The intraspecific genetic distance was smaller than the interspecific ones;The NJ tree indicated that Pinelliae Rhizoma distinguished from its adulterants obviously. The results showed that the ITS2 sequence can be used to distinguish Pinelliae Rhizoma from its adulterants accurately and stably.
ABSTRACT
Objective: This study aimed to discriminate between Eupatorii Herba and its adulterants in order to guarantee the quality and clinical curative effect of this medicinal material. Methods: Genomic DNA extracted from Eupatorii Herba was used as templates. The internal transcribed spacer 2 (ITS2) of nuclear ribosomal DNA was amplified. Sequence assembly and consensus sequence generation were performed by CodonCode Aligner. The intraspecific and interspecific genetic distances of Eupatorii Herba and its adulterants were computed by MEGA5 and the phylogenetic tree was constructed using the neighbor-joining (NJ) method. Results: The length of ITS2 sequence of Eupatorii Herba was 218 bp. The maximum intraspecific genetic distance (K2P distance) of Eupatorii Herba was 0.0092. The minimum interspecific genetic distance of Eupatorii Herba and its adulterants was 0.024. The NJ trees showed that the ITS2 sequence would be used to identify Eupatorii Herba and its adulterants. Con-clusion: ITS2 sequence was able to identify Eupatorii Herba and its adulterants correctly and it provided a new technique to ensure clinical safety in utilization of traditional Chinese medicines.