Varieties Classification Of Osmanthus Fragrans Lour. Using Random Amplified Polymorphic DNA (RAPD)

This paper aimed at the analysis on variety classification for Osmanthus fragrans (Thunb.) Lour. using RAPD (Random Amplified Polymorphic DNA) method. The 80 selected plants belong to four variety groups, namely Latifolius group, Thunbergii group, Autiacus group, Fragrans group. The selection of amplified reaction program and system was carried out. 20 primers for RAPD analysis were obtained during the first selection, 9 for second selection, which could be used for UMPGA (Unweighted pair-group mathematic analysis). The results were as follows:1 The best way selected for Osmanthus fragrans (Thunb.) Lour. DNA exacting was modified CTAB method.2 The Amplified reaction system for RAPD analysis was: total amount 25 μl; 2μl primer (5pmol/μl) + template DNA 40ng+2.5μl PCR buffer( 500mmol/LTris-Cl, pH8.3, 5.00mmol/L KCl,15mmol/L MgCl₂, 0.01% Gelatin) + 1 .0UTaqDNApolymerase + 0. 4μl100mmol/LdNTPs(each 25mmol/L) .The mixture was joined with super-pure water .3 The Amplified reaction program was : 94℃, initial denaturation 3 min→94℃50s → 36℃ 1min→72℃ 5min, 38 times repeat→72℃ 10min, stored at 4℃ 12 hr.4 The screening of 80 dreamer oligoncleotides of OP system resulting in the selection of 20 primers suiting for RAPD analysis for Osmanthus fragrans Lour. . 9 primers were selected again from the 20 primers to analysis the samples in the sequent work.5 The result of polymorphism analysis: There were a total of 47 bands, and 45 bands were polymorphic. The polymorphic rate reached to 95.7%. According to the result of the analysis of polymorphism, the 80 samples were showed a complex genetic base. The clustering results according to genetic similarity coefficients and genetic distance were not in consistence with that of the analysis of morphology.6 The result of variety classification: there was a short genetic distance in the same variety group, and a long one between different variety groups. The clustering results according to genetic similarity coefficients and genetic distance were not in consistence with that of the analysis of morphology.