Abstract
Salvia miltiorrhiza (SM), a widely popular Chinese herb, is grown in various regions in China. Identifying SMs grown in different provinces of China is difficult, and therefore genotyping these collections would be highly valuable. Based on the techniques of sequence-related amplified polymorphism and target region amplified polymorphism, a novel PCR-based molecular marker technique called conserved region amplification polymorphism (CoRAP) is reported in this study to genotype SMs. The CoRAP technique is based on the use of two primers: fixed and arbitrary primers. The former is derived from target EST sequences deposited in Genbank; while, the core sequence (CACGC) of the latter is a conserved region found in most introns. In the present study, we utilized CoRAP to genotype SMs from different geographical origins. PCR amplification is performed for 30 cycles at an annealing temperature of 52°C. Each PCR reaction has generated as many as 30–50 fragments of 50 to 1,000 bp in size. The successful DNA genotyping of SMs by CoRAP was achieved. This new genotyping method is rapid, efficient, and reproducible.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
Abbreviations
- CoRAP:
-
conserved region amplification polymorphism
- EST:
-
expressed sequence tag
- PCR:
-
polymerase chain reaction
- SM:
-
Salvia miltiorrhiza
References
Abd-Elazem IS, Chen HS, Bates RB, Huang RC. Isolation of two highly potent and non-toxic inhibitors of human immunodeficiency virus type 1 (HIV-1) integrase from Salvia miltiorrhiza. Antiviral Res. 2002;55:91–106. doi:10.1016/S0166-3542(02)00011-6.
Botstein D, White RL, Skolnick M, Davis RW. Construction of a genetic linkage map in man using restriction fragment length polymorphisms. Am J Hum Genet. 1980;32:314–31.
Cheng TO. Cardiovascular effects of Danshen. Int J Cardiol. 2007;121:9–22. doi:10.1016/j.ijcard.2007.01.004.
Hsu YC, Lin YL, Chiu YT, et al. Antifibrotic effects of Salvia miltiorrhiza on dimethylnitrosamine-intoxicated rats. J Biomed Sci. 2005;12:185–95. doi:10.1007/s11373-004-8167-7.
Hu J, Vick BA. Target region amplification polymorphism: a novel marker technique for plant genotyping. Plant Mol Biol Rep. 2003;21:289–94. doi:10.1007/BF02772804.
Janssen P, Coopman R, Huys G, Swings J, Bleeker M, Vos P, et al. Evaluation of the DNA fingerprinting method AFLP as an new tool in bacterial taxonomy. Microbiology 1996;142(Pt 7):1881–93.
Li G, Quiros CF. Sequence-related amplified polymorphism (SRAP), a new marker system based on a simple PCR reaction: its application to mapping and gene tagging in Brassica. Theor Appl Genet. 2001;103:455–61. doi:10.1007/s001220100570.
Litt M, Luty JA. A hypervariable microsatellite revealed by in vitro amplification of a dinucleotide repeat within the cardiac muscle actin gene. Am J Hum Genet. 1989;44:397–401.
Moore SS, Sargeant LL, King TJ, Mattick JS, Georges M, Hetzel DJ. The conservation of dinucleotide microsatellites among mammalian genomes allows the use of heterologous PCR primer pairs in closely related species. Genomics 1991;10:654–60. doi:10.1016/0888-7543(91)90448-N.
Tautz D. Hypervariability of simple sequences as a general source for polymorphic DNA markers. Nucleic Acids Res. 1989;17:6463–71. doi:10.1093/nar/17.16.6463.
Vos P, Hogers R, Bleeker M, Reijans M, van de Lee T, Hornes M, et al. AFLP: a new technique for DNA finger printing. Nucleic Acids Res. 1995;23:4407–14. doi:10.1093/nar/23.21.4407.
Vuylsteke M, Peleman JD, van Eijk MJ. AFLP technology for DNA fingerprinting. Nat Protocols. 2007;2:1387–98. doi:10.1038/nprot.2007.175.
Welsh J, McClelland M. Fingerprinting genomes using PCR with arbitrary primers. Nucleic Acids Res. 1990;18:7213–8. doi:10.1093/nar/18.24.7213.
Williams JG, Kubelik AR, Livak KJ, Rafalski JA, Tingey SV. DNA polymorphisms amplified by arbitrary primers are useful as genetic markers. Nucleic Acids Res. 1990;18:6531–5. doi:10.1093/nar/18.22.6531.
Zhou L, Zuo Z, Chow MS. Danshen: an overview of its chemistry, pharmacology, pharmacokinetics, and clinical use. J Clin Pharmacol. 2005;45:1345–59. doi:10.1177/0091270005282630.
Acknowledgements
These studies were supported, in part, by the National Natural Science Foundation of China grant, 30672613.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Wang, Q., Zhang, B. & Lu, Q. Conserved Region Amplification Polymorphism (CoRAP), a Novel Marker Technique for Plant Genotyping in Salvia miltiorrhiza . Plant Mol Biol Rep 27, 139–143 (2009). https://doi.org/10.1007/s11105-008-0065-0
Published:
Issue Date:
DOI: https://doi.org/10.1007/s11105-008-0065-0