Identification of random amplified polymorphic DNA (RAPD) marker for differentiating male from female and sporophytic thalli of Gracilaria changii (Rhodophyta)
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There have been limited reports on molecular sex markers for macroalgae. We report the use of random amplified polymorphic DNA analysis (RAPD) to identify molecular sex markers for Gracilaria changii (Xia et Abbott) Abbott, Zhang et Xia. Two DNA extraction methods were used: a modified CTAB and phenol-chloroform combination method and the DNeasy Plant Mini Kit. The CTAB and phenol-chloroform method gave the best yield of DNA in quality and quantity and is suitable for larger-sized specimens like G. changii. Sixty-nine RAPD primers were screened to search for sex-linked DNA markers for G. changii, and only one sex-linked marker (716 bp) was identified using OPA 18. RAPD was also used to investigate the molecular characteristics of the three life-stages (male, female, tetrasporophyte) of G. changii. Seven (OPA7, OPA18, S14, S61, S64, S75 and S76) out of the 69 primers showed polymorphism and were selected for interpopulation analysis for DNA isolated from 23 samples collected from Morib and Sungai Pulai in Malaysia. The combination of data produced by the seven primers generated a dendrogram that grouped the specimens into different clades according to their sex and life-stage using the unweighted pair group and arithmetic averages (UPGMA) method. It showed that RAPD was able to differentiate tetrasporophytes, females, and males.
KeywordsGracilaria changii Molecular sex markers Random amplified polymorphic DNA (RAPD)
The authors gratefully acknowledge the following: Ministry of Science, Technology and Innovation, Malaysia (IRPA Grant Nos. 39-02-03-9002), and Vote F (0187/2004B), University of Malaya, for providing financial assistance and Professor Kwai Lin Thong for her advice and the use of her equipment.
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