Molecular Authentication of Scrophularia herbs by PCR-RFLP Based on rpl-5 Region of Mitochondrial DNA

현삼속 식물의 종판별을 위한 Mitochondrial DNA의 염기서열 및 PCR-RFLP 분석

  • 이정훈 (국립원예특작과학원 인삼특작부) ;
  • 조익현 (국립원예특작과학원 인삼특작부) ;
  • 이제완 (국립원예특작과학원 인삼특작부) ;
  • 박춘근 (국립원예특작과학원 인삼특작부) ;
  • 방경환 (국립원예특작과학원 인삼특작부) ;
  • 김홍식 (충북대학교 식물자원학과) ;
  • 박충범 (국립원예특작과학원 인삼특작부)
  • Received : 2010.05.12
  • Accepted : 2010.06.11
  • Published : 2010.06.30

Abstract

This study describes an efficient approach to the development of DNA markers for use in distinguishing the Scrophularia species that have been used as useful medicinal crops. In order to distinguish Scrophularia species, DNA sequences of rpl-5 region in mitochondrial DNA of Scrophularia species were analysed for detecting sequence variations, and the PCR-RFLP method was applied for developing practicable DNA marker patterns. Several DNA variations were detected by the sequence comparison of rpl-5 region among Scrophularia species. Genetic relationship analysis of Scrophularia species was carried out based on these DNA variations. DNA variations of rpl-5 region were revealed that it was significantly efficient in genetic relationship analysis of Scrophularia species. In addition, Scrophularia species tested in this study were completely discriminated by four polymorphic genotypes by PCR-RFLP combined with Tsp509 I (^AATT) restriction enzyme. Our results suggested that DNA sequence variations of rpl-5 region were sufficiently useful for genetic relationship analysis of Scrophularia species. Polymorphic genotypes by PCR-RFLP using the Tsp509 I enzyme will be useful for discrimination of Scrophularia species as a practicable DNA markers.

Keywords

References

  1. Chae YA, Kim SM, Kim KS, Yu CY, Yang DC, Song CK and Kang JH. (2007). Medicinal Phytology. Hyangmoonsa Press. Seoul, Korea. p. 223-224.
  2. Duminil J, Pemonge MH and Petit RJ. (2002). A set of 35 consensus primer pair amplifying gene and introns of plant mitochondrial DNA. Molecular Ecology Note. 2:428-430. https://doi.org/10.1046/j.1471-8286.2002.00263.x
  3. Felsenstein J. (1985). Confidence limits on phylogenies: An approach using the bootstrap. Evolution. 39:783-791. https://doi.org/10.2307/2408678
  4. Han KS, So SK, Lee CH and Kim MY. (2009). Taxonomy of the genus Scrophularia (Scrophulariaceae) in Korea. The Korean Journal of Plant Taxonomy. 39:237-246. https://doi.org/10.11110/kjpt.2009.39.4.237
  5. Hwang SH, Byun JH, Song HJ and Shin MK. (1998). A herbalogical study on the plants of Scrophulariaceae in Korea. Journal of the Herbology Society of Korea. 13:241-270.
  6. Jigden B, Kim MK, Noh JH, Hua S and Yang DC. (2009). Phylogenetic analysis of Schizonepeta spike on the basis of DNA sequences. Korean Journal of Medicinal Crop Science. 17:46-53.
  7. Kim EJ, Kim SR, Kim J and Kim YC. (2002). Hepatoprotective phenylpropanoids from Scrophularia buergeriana roots against CCl4-induced toxicity: action mechanism and structure-activity relationship. Planta Medica. 68:407-411. https://doi.org/10.1055/s-2002-32081
  8. Kim MK, Jigden B, Hua S, Noh JH, Kim SY and Yang DC. (2008). Phylogenetic analysis of Ji-Mo (Anemarrhena asphodeloides) on the basis of the chloroplast DNA sequences. Korean Journal of Medicinal Crop Science. 16:20-26.
  9. Kim SR and Kim YC. (2000). Neuroprotactive phenylpropanoid esters of rhamnose isolated from roots do Scrophularia buergeriana. Phytochemistry. 54:503-509. https://doi.org/10.1016/S0031-9422(00)00110-2
  10. Kim SR, Sung SH, Jang YP, Markelonis George J, Oh TH and Kim YC. (2002). E--Methoxycinnamic acid protects cultured neuronal cells against neurotoxicity induced by glutamate. British Journal of Pharmacology. 135:1281-1291. https://doi.org/10.1038/sj.bjp.0704576
  11. Kim SR, Kang SY, Lee KY, Kim SH, Markelonis George J, Oh TH and Kim YC. (2003). Anti-amnestic activity of E-pmethoxycinnamic acid from Scrophularia buergeriana. Cognitive Brain Research. 17:454-461. https://doi.org/10.1016/S0926-6410(03)00161-7
  12. Lee HW, Park SY, Choo BK, Chun JM, Lee AY and Kim HK. (2007). Quantitative analysis comparison of Korea and China Scrophulariae radix. Korean Journal of Pharmcognosy. 38:15- 18.
  13. Lee JH, Lee JW, Sung JS, Bang KH and Moon SK. (2009). Molecular authentication of 21 Korean Artemisia species (Compositae) by polymerase chain reaction-restriction fragment length polymorphism based on trnL-F region of chloroplast DNA. Biological and Pharmaceutical Bulletin. 32:1912-1916. https://doi.org/10.1248/bpb.32.1912
  14. Lee YJ and Ahn DK. (1986). The study of Scrophularia (Scrophulariaceae) about origin and effect. Journal of the Herbology Society of Korea. 1:63-72.
  15. Park KH, Ham IH and Choi HY. (2008). Hypotensive effect of Scrophularia buergeriana and gene analysis of Scrophularia species. Journal of the Herbology Society of Korea. 23:213- 223.
  16. Schuster W. (1993). Ribosomal protein gene rpl 5 is cotranscribed with the nad 3 gene in Oenothera mitochondria. Molecular and General Genetics. 240:445-449.
  17. Song JS, Lim WS and Chae YA. (1998). Effects of explants and growth regulators on direct somatic embryogenesis in liquid culture of Scrophularia buergeriana. Korean Journal of Medicinal Crop Science. 6:294-298.
  18. Tamura K, Dudley J, Nei M and Kumar S. (2007). MEGA4: molecular evolutionary genetic analysis (MEGA) software version 4.0. Molecular Biology and Evolution. 24:1596-1599. https://doi.org/10.1093/molbev/msm092
  19. Vincze T, Torrell M, Garnatje T, Garcia-Jacas N, Vilatersana R and Susanna A. (2003). NEB cutter: a program to cleave DNA with restriction enzyme. Nucleic Acids Research. 31:3688-3691. https://doi.org/10.1093/nar/gkg526
  20. Woo WS. (1963). Active principle, ρ-Methoxycinnamic acid, of scrophulariae radix . identification of ρ-Methoxycinnamic acid and its antipyretic action. Korean Journal of Pharmcognosy. 7:55-57.
  21. Woo WS. (1965). Analgesic action of $\rho$-Methoxycinnamic acid. Korean Journal of Pharmcognosy. 9:31-33.
  22. Yoon IS, Song HJ, Kim SJ, Kang DH, Kim JM, Choi GY and Ju YS. (2007). A herbological study on the herbs in 3 kinds of common Scrophulariaceae. The Journal of Korean Oriental Medical Ophthalmology & Otolaryngology & Dermatology. 20:127-137.
  23. Yu CY, Seong ES, Lim JD, Huang SA and Chae YA. (2001). Introduction and expression of PAP gene using Agrobacterium in Scrophularia buergeriana Miquel. Korean Journal of Medicinal Crop Science. 9:156-165.