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Abstract Rubber is a kind of important strategic material in China. There are important significance in rubber tree genomic study. In this study, the chromosome samples were prepared with the enzyme and low osmotic method using tender leaves of Hevea brasiliensis RY-7-33-97 as material. The preparation of chromosome was optimized. Chromosomes were successfully isolated by slender glass needles microdissection. The microdissected chromosomes was digested with proteinase K, and its DNA was extracted and digested with Sau3A, amplified by linker adaptormediated PCR(LA-PCR) for 2 rounds, a smear of DNA fragments ranging from 250 to 2 000 bp was generated. Amplified single chromosome production was verified by Southern blot. The results showed that the LA-PCR products were homogeneous with the H. brasiliensis genomic DNA, indicating that DNAs from the chromosome had been successfully amplified. The ninth chromosome was verified by fluorescence in situ hybridization (FISH), and constructed microcloning library of the ninth chromosome in H. brasiliensis. The library contains 48 000 clones. The inserts averaged 650 bp, and varied from 300 to 1 200 bp with no-load rate of 1%. In this study, construction of the single chromosome library of rubber tree, by the means of single chromosome microdissection and microcloning technology, will provide a new perspective for the study of rubber tree genome and a molecular assisting way for breeding of rubber tree.
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Received: 03 March 2014
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