PertanikaJ. Trap. Agric. Sci. 22(2): 111-116 (1999) ISSN: 1511-3701© Universiti Putra Malaysia Press

A Preliminary Study of Genetic Variation of Selected Species of a LowlandForest at Ayer Hitam Forest Reserve, Selangor

OR AI I AB. SHUKORFaculty of Forestry

Universiti Putra Malaysia43400 UPM Serdang, Selangor, Malaysia

Keywords : genetic variation, starch gel electrophoresis, polymorphism, heterozygosities

ABSTRAK

Kajian awal mengenai corak variasi genetik ke atas 10 spesies pilihan (Shorea parvifolia, Shorea macroptera,Shorea acuminata, Shorea leprosula, Hopea beccariana, Dipterocarpus crinitus, Endospermum malaccensis,Artocarpus elasticus, Palaquium gutta dan Macaranga gigantea) dari Hutan Simpan Ayer Hitam telahdijalankan menggunakan kaedah elektroforesis gel kanji. Analisis ke atas 8 enzim menunjukkan yang rnerekadikawal oleh 9 ke 12 lokus. Tahap polimorfik dan purata heterozigositi jangkaan bagi spesies-spesies ini berjulatdaripada 0.9 ke 1.0 dan daripada 0.454 (S. leprosula) ke 0.602 (H. beccariana) masing-masing.

ABSTRACT

A preliminary study of the extent and pattern ofgenetic variation of 10 selected species (Shorea parvifolia, Shoreamacroptera, Shorea acuminata, Shorea leprosula, Hopea beccariana, Dipterocarpus crinitus, Endospermummalaccensis, Artocarpus elasticus, Palaquium gutta and Macaranga gigantea) at Ayer Hitam Forest Reserve wascarried out using the horizontal starch gel electrophoresis. Analysis ofeight enzymes indicated that they were codedlJy 9 to 12 loci. Levels of polymorphism and mean of expected heterozygosities of these species ranged from 0.9to 1.0 and from 0.454 (S. leprosula) to 0.602 (H. beccariana) respectively.

INTRODUCTION

Tropical forest is rich in genetic resources. Highspecies diversity is the most peculiar feature oftropical rainforest. For instance, the dipterocarpsare the most dominant canopy component andrepresent the richness of tropical taxa (Ashton1988). The complex community process andreproductive process of these diversified speciesare challenged by recent tropical ecology (Ashton1988). Although the genetic studies of tropicalspecies are expected to provide importantinformation on the community, organisation andreproductive ecology, studies on native treespecies in tropics are relatively scarce (Hamrickand Loveless, 1986; Loveless 1992 and Kijkar1992) .

Effective forest management needs athorough understanding of many aspectsincluding biology, ecology and genetics.Information on the genetics of species would beuseful in designing appropriate tree breeding

programmes. Lewontin (1974), Endler (1977)and Loveless (1992) stated that the presence ofvariability within a population is responsible ingenerating and maintaining a populationsustainably. Genetic variation is the fundamentalrequirement for the maintenance and long termstability of forest ecosystem since the amountand pattern of genetic variation would determinethe ability of forest tree species to adapt tovariable environmental conditions (Bergmann etal. 1989). In fact, many studies of geneticvariation showed that it is correlated with lifehistory characteristics, breeding system andpopulation dynamics (Loveless and Hamrick1984). Genetic studies can also help in theidentification of superior populations.

Among the many reliable and practicalmethods used in the study of genetic variation isby isozyme analysis. Isozyme markers have beenfound to be the cheapest and reliable whencompared to the traditional morphological

OR AlNI AB. SHUKOR

markers. Its utilisation enables the separation ofproducts of the same genes, regardless ofenvironment. Individuals may be characterised bytheir genotypes, composed of a sample of gene,comparison of individuals or groups of individualscould be made using specific genetic markers.

Thus, the objective of this study is to assessthe genetic variation (inter and intra specific) of10 selected lowland forest species at Ayer HitamForest Reserve, Selangor.

MATERIALS AND METHODS

Samples of cambial tissues of 10 selected species(Shorea parvifolia, Shorea macroptera, Shoreaacuminata, Shorea leprosula, Hopea beccariana,Dipterocarpus crinitus, Endospermum malaccensis,Artocarpus elasticus, Palaquium gutta andMacaranga gigantea) were collected fromcompartment 15, Ayer Hitam Forest Reserve,Selangor. Cambial tissues were collected from30 trees per species to determine the totalvariation of the species. The samples were keptin eppendorf tubes and placed in a cooling boxduring the transportation. They were thenfiltered and mixed with extraction buffer priorto electrophoretic run.

The buffers used in this study were basedon the ones recommended by previous studieson Shorea species (Daisy 1995 and Noridah, 1996).The three buffers used were Histidine (H) ,Lithium (L) and Morpholine Citrate (Me).Electrophoresis was done on a 10.5% potatohydrolysed starch. Mter electrophoresis, the gelswere stained for 8 different enzymes namelyEsterase (EST), a Glycerophosphate dehydrogenase(aCPDH), Isocitrate dehydrogenase (lDH), Malatedehydrogenase (MDH), Phosphoglucomutase (PCM) ,6- Phosphoglucose Dehydrogenase (6 PDCH),Phosphoglucose isomerase (PCI) and ShikimateDehydrogenase (SDH). The genetic interpretationof the enzyme was based on the phenotypesobtained according to the mobility of the isozymebands. When an enzyme revealed more thanone zone of activity, the fastest migrating (mostanodal) was designated as locus 1, the next, 2and so on. In addition, the most anodal allelewas labelled as fast (F) , medium (M) and theleast migrated allele as slow (S). Clear bandingpatterns were obtained for all 9 loci and theresults of the electrophoretic phenotype variationwere analysed for allelic frequencies andexpected heterozygosities.

RESULTS AND DISCUSSION

Interpretation of the enzyme phenotypes wasbased on patterns of variability in 10 species.Direct verification of genetic control of theelectrophoretic patterns of the enzymesexamined was not carried out. It was assumedbased on the consistency of the electrophoreticpatterns of the monomorphic and polymorphicenzymes from each individual of the speciesanalysed. A total of 9-12 loci were scored fromthe eight enzyme systems. Almost all of the lociscored were polymorphic except for Sdh-l in S.acuminata and Pgi-2 for P. gutta. The averagenumber of alleles per locus ranged from 2.4 to2.9 (Table 1).

The interspecific genetic variation isquantified by measuring the mean heterozygosity,the percentage of polymorphic loci and thepercentage of alleles per locus. The meanobserved heterozygosities (Ho) of these speciesranged from 0.383 in S. acuminata to 0.608 in P.gutta while the mean of expected heterozygositiesranged from 0.454 to 0.602. On the other hand,level of polymorphic loci ranged from 0.9 to 1.0(Table 1). Values of heterozygosities andpolyrnorphisms were found to be similar to thosereported by Kong (1994) on Shorea acuminata(Ho=0.604, P=l.O); Daim (1993) and Daisy (1995)on S. leprosula (Ho=0.565, and Ho=0.457, P=0.9respectively) (Table 2). However, these valueswere higher than the values given by Hamrickand Loveless (1986), Loveless and Hamrick(1987), John (1996) and Hazandy (1997) forother tropical tree species (Table 2).

The higher values of heterozygosities andpolymorphism in this study indicated that thesespecies have undergone effective competition andselection within and between species for survivalin natural stand. The natural forest, whichnormally portrays heterogeneous ecologicalcondition, would justify them to possess highgenetic variability. Thus, the heterozygotes beingthe fitter genotypes would survive better due tothese effects as being indicated by the highervalues of Ho over He (Feret and Bergmann,1976) (Table 1). Hamrick and Loveless (1984)also found that the genetic variation of treespecies in natural stand higher almost doublethat of other plant species. In addition, Hamrick(1989) and Hamrick et al. (1992) also reportedthat long-lived woody perennials would show highlevels of genetic diversity. Hamrick and Codt(1989) and Hamrick et al. (1992) showed that He

112 PERT lKAJ. TROP. ACRIC. SCI. VOL. 22 0.2,1999

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TABLE 1Summary on the Mean Observed Heterozygosity (H), Mean Expected

Heterozygozity (H) and Proportion of Polymorphic Loci.

S. S. S. S. H. D. E. A. P. M.parvifolia macroptera acuminata leprosula beccariana crinitus malaccensis elasticus gutta gigantea

(30) (30) (30) (30) (30) (30) (30) (30) (30) (30)

RangeH

00.000 0.200 0.000 0.000 0.100 0.200 0.000 0.100 0.300 0.100

H0

0.800 1.000 0.900 0.800 1.000 0.900 0.900 0.900 1.000 0.800Mean 0.446 0.562 0.383 0.510 0.473 0.467 0.422 0.558 0.608 0.440RangeH e 0.255 0.255 0.420 0.255 0.445 0.180 0.322 0.460 0.000 0.255H e 0.825 0.743 0.990 0.620 0.695 0.906 0.655 0.665 0.640 0.790Mean 0.512 0.506 0.582 0.454 0.602 0.471 0.536 0.509 0.482 0.537L 11 12 12 10 11 9 9 12 12 10

P 1.0 1.0 0.9 1.0 1.0 1.0 1.0 1.0 0.9 1.0

A 2.7 2.6 2.4 2.5 2.9 2.6 2.7 2.8 2.6 2.7

Note: Number in parentheses indicate sample size.Key : H

o- observed heterozygosity, He - expected heterozygosity;

L - loci scoredP - proportion of polymorphic lociA - average number of alleles per locus

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NOR AINI AB. SHUKOR

TABLE 2Genetic diversity parameters of tropical tree species as compared to genetic

diversity of ten species obtained in this study.

Species

Acacia auriculiformisA. auriculiformisA. crassicarpaA. crassicarpaA. mangiumArtocarpus elasticusAzadirachta excelsaDipterocarpus crinitusEndospermum malacensisGliricidia sepiumH evea brasiliensisHopea beccarianaH opea odorataLeucaena shannonniiMacaranga giganteaPalaquium guttaPinus kesiyaPterocarpus macrocarpusShorea acuminataS. acuminataS. leprosulaS. leprosulaS. macrophyllaS. macropteraS. leprosulaS. parvifoliaS. parvifoliaTectona grandisTropical species

Note: n.a. - not available.

HeterozygosityExpected

0.0710.1460.0860.1410.2000.5080.0940.4710.5360.2600.3070.6020.1900.2710.5380.4820.1660.2460.6040.5820.5650.4570.2090.5070.4540.5350.5120.3470.111

Proportion ofPolymorphic

Loci (%)

39.8n.a.58.7

n.a.67.0

100.067.5

100.0100.0

59.987.5

100.040.765.7

100.091.654.282.3

100.091.772.293.347.0

100.0100.0

95.2100.0

79.027.6

References

Wickneswari and Norwati (1993)Moran et al. (1989a)John (1996)Moran et al. (1989a)Hamidi (1990)Present studyHazandy (1997)Present studyPresent studyChamberlain et al. (1996a)de Paiva et al. (1994)Present studyWickneswari et al. (1995)Chamberlain et al. (1996b)Present studyPresent studyBoyle et al. (1991)Liengsiri et al. (1995)Kong (1994)Present studyDa im (1993)Daisy (1995)Kanzaki et al. (1996)Present studyPresent studyKong (1994)Present studyKertadikara and Prat (1995)Hamrick and Loveless (1986)

depends on the geographic range of the speciesi.e. species with wider distribution range hashigher He' The outcrossing nature of most ofthese species can further support such result. Anoutcrossed species has been reported to producea more genetically diverse sample (Moran et al.1989a), especially when samples were taken fromindividuals originating possibly from only a fewmother trees. This phenomenon would be likelyto occur for species that experience prevalent asexual reproduction i.e. by apomixis. For instanceKaur et al. (1978);(1986) and Somego (1978)have provided some evidence of apomixis ofsome Malaysian dipterocarps. Thus detailedreproductive biology and cytological studiesshould be incorporated to clarify the observedgenetic status of these species.

CONCLUSION

Genetic variation among the selected species isrelatively high in terms of polymorphism andheterozygosity values. High levels of geneticvariability for all species are strongly associatedwith the life history, reproductive biology andtheir capability for adaptation. The averagegenetic variation of the available species with Heof 0.454 to 0.602 is high and sufficient to supporta selective breeding programme. However,further verification of genetic control of enzymeloci assayed is required. In addition, evaluationof these parameters should be conducted onsimilar species from different compartments toassess for the intraspecific variation before anysampling and breeding strategies could beoutlined. In addition, growth performances of

114 PERTANlKAJ. TRap. AGRIC. SCI. VOL. 22 NO.2, 1999

A PRELIMINARY STUDY OF GENETIC VARIATION OF SELECTED SPECIES OF A LOWlAND FOREST

these species should also be evaluated so as tocapture maximum genetic gain for the purposesof breeding and conservation.

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