Generation of SCAR Markers for Identification of Satellite Chromosome from Cunninghamia lanceolata Seeds

Abstract:

To generate markers of sequence characterized amplified regions (SCAR), we recovered and purified a fragment OPB07-18907 from satellite chromosome of Cunninghamia lanceolata (Lamb.) Hook, using random amplified polymorphic DNA (RAPD) pairwise primers OPB07(5’-GGTGACGCAG-3’) and OPB18（5’-CCACAGCAGT-3’）. This fragment was then cloned into pUCm-T vector，and transferred into E. coli. DH5α. Positive colonies were identified for sequencing. Two SCAR primer pairs were designed according to the sequences of OPB07-18₉₀₇. Based on information of the fragment, the SCAR marker was produced for satellite chromosome. The products of SCAR-PCR indicated that all primer combinations could be used to amplify specific bands of satellite chromosome. In addition, we found that 57℃ was the optimal annealing temperature for the use of these primers. We developed the SCAR marker of satellite chromosome for identifying the microdissected satellite chromosome at molecular level, and correlating a genetic linkage group of C. lanceolata with a specific chromosome. We also discussed the implementation of SCAR marker in karyotype polymorphism in C. lanceolata.