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Assessment of Genetic Diversity in Garcinia Species by using Isozyme Analysis, Random Amplified Polymorphic DNA (RAPD), and Simple Sequence Repeats (SSRs)
Thesis Abstract:
Genetic diversity in 22 accessions of Garcinia species was assessed using isozyme analysis, random amplified polymorphic DNA (RAPD), and simple sequence repeats (SSRs).
Among the 15 isozymes tested, only peroxidase isozyme produced bands. A total of eight bands were generated with relative mobility (Rf) value between 0.57 and 0.75 forming three peroxidase isozyme patterns distinct for G. mangostana, G. binucao, G. kydia, and G. lateriflora. No band was observed in G. livingstonei and G. xanthochymus.
RAPDs and SSRs were used to confirm the isozyme results. The extraction of genomic DNA of the different Garcinia species was done using Cheung, Hubert, and Landry (1993)* buffer. For RAPDs, the extracted DNA was subjected to polymerase chain reaction (PCR) condition using the operon primers (OPB-04, OPB-06, and OPB-07). For SSR, two sets of SSR primers were designed based on each consensus nucleotide sequence of the two enzymes expressed in Garcinia sp. primer acyl-acyl carrier protein (ACP) for acyl-ACP thioesterase, and primer CHALCS for chalcone synthase using Primer3 program. The conserved flanking regions were identified as sites for primer design using Vector NTI program suite. The polymorphism information content is 0.839 and 0.842 for acyl-ACP and CHALCS, respectively.
The higher level of informativeness by the SSR markers indicated the capability of microsatellites to quantify genetic diversity in Garcinia species. The proportion of shared alleles from three operon primers and nine alleles detected from SSR markers derived from acyl-ACP synthase and chalcone synthase for each locus revealed five banding patterns.
Underweight pair group method with arithmetic mean (UPGMA)-sequential, agglomerative, hierarchical, and non-overlapping (SAHN) cluster analysis of RAPDs and SSRs data revealed similar pattern of grouping. Dendogram for RAPDs and SSRs showed that Garcinia species clustered into five at coefficient 0.330 and 0.502, respectively. The RAPDs and SSRs results revealed that group 1 was composed of all 17 G. mangostana accessions. Group 2 was composed of G. kydia and G. lateriflora. Group 3 was composed of individuals of G. livingstonei, G. binucao, and G. xanthochymus.
This study showed that all mangosteen accessions have the same genetic origin. Further, results also revealed that there was misidentification of either G. kydia or G. lateriflora in the samples studied.
∗ Χηευνγ, Ν.Ψ., Ν. Ηυβερτ, ανδ Β.ϑ. Λανδρψ. 1993. Α Σιμπλε ανδ Ραπιδ ΔΝΑ Μιχροεξτενσιον Μετηοδ φορ Πλαντ, Ανιμαλ ανδ Ινσεχτσ Συιταβλε φορ ΡΑΠΔ αφτερ ΠΧΡ αναλψσεσ. ΠΧΡ Μετσ Αππλ. 3:69−70.