Development and characterization of chloroplast microsatellite markers for Pinus massoniana and their application in Pinus (Pinaceae) species

Pinus massoniana is one of the important afforestation and pioneer tree species, which is widely distribute in southern China. Chloroplast simple sequence repeat markers (cpSSRs) have been widely used in studies of tree genetics, phylogenetic and breeding. We sequenced the whole chloroplast genome sequences of P. massoniana using PCR and Sanger sequencing. A total of 71 cpSSRs were identified, among which mononucleotide repeats were predominant (70.42%). Seventeen primer pairs were developed and amplification tests were conducted with 15 P. massoniana individuals. Also, cross-species amplification tests were conducted among 15 individuals per Pinus species, including P. elliottii, P. bungeana, P. armandii, P. caribaea, P. tabulaeformis, P. taiwanensis and P. yunnanensis which revealed polymorphic information content ranging from 0.2 to 0.8 and average of haploid diversity (h) ranging from 0.29 to 0.63. In addition, the polymorphic cpSSRs were useful in distinguishing the sampled pine species, and could be powerful tool in phylogenetic studies.