Molecular cloning and sequencing of a polymorphic band from rubber tree (Hevea brasiliensis (Muell.) Arg.): the nucleotide sequence revealed partial homology with proline-specific permease gene sequence †

Rubber tree (Hevea brasiliensis Muell. Arg.) is mainly cultivated in high rainfall areas (traditional). Identification of DNA marker or genes could be highly useful to develop drought-tolerant clones or screen Hevea seedling populations, which will eventually extend the rubber cultivation in non-traditional areas (under irrigation). RAPD analysis is a valuable tool in studying genetic variation and for identification of specific gene fragments. Genomic DNAs from 37 clones were used for RAPD analysis by polymerase chain reaction (PCR), where the total and unique number of polymorphic bands varied with the clones and primers. Certain amplified bands appeared to be common to some clones, whereas others were present only in a few clones. RAPD markers specific to one or few clones were identified, with the results being reproduced at least three times. There were two DNA markers (1.2 and 1.4 kb) scored by presence vs absence of specific amplification products with OPB-12 primer. Of the two bands, a 1.4 kb RAPD marker was the most promising DNA fragment which was isolated, cloned and the nucleotide sequence determined and revealed certain homology with Saccharomyces cerevisiae proline-specific permease gene. Specific DNA technology, such as the use of SCAR (sequence characterized amplified region) markers, can be used to differentiate among Hevea clones to validate this RAPD marker. SCAR marker was created by sequencing a single RAPD band and designing primers to amplify the band of specific size. The SCAR primers amplify a band of 1.4 kb from only 14 among 37 clones used under more stringent PCR conditions. This SCAR marker could be used as genetic marker to screen the Hevea seedling populations for specific trait or gene from single PCR-based method.