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CONSERVATION STRATEGIES OF SHOREA LUMUTENSIS (DIPTEROCARPACEAE) IN PENINSULAR MALAYSIA
regeneration (Condit et al. 1998). Short-term population dynamics was derived from the initial
census in September 2001 and a repeat census in August 2004 for growth rates (based on
increase in dbh) and mortality.
Flowering and Fruiting Biology
Phenological observations were carried out using binocular from January 2002 to October
2005. Periodic surveillance was undertaken following Appanah & Chan (1982) to establish
the flowering stages (budding, initial bloom, peak bloom, tail bloom, and termination of bloom),
flowering intensity (intense, moderate, and poor) and fruiting stages (seed development, seed
maturing and seed fall).
Germination and Seedling Studies
Seed collections were conducted in December 2002 for trees B026 and B385, and in January
2003 for trees B004 and B005 within the 8-ha study plot in Sungai Pinang. The seed weight
variation and its effect on germination and speed of germination were tested using binary and
ordinal logistic regression analyses, respectively. The relationship between seed weight and
seedling vigor (seedling height after three months of growth) was tested for y = bx + c, in
which y represents the seedling height, x the seed weight, and a and b are the intercept and the
slope of the curve, respectively.
Development of Microsatellite Loci
The total genomic DNA was extracted from leaf tissues using the procedure described by
Murray & Thompson (1980), with modification, and further purified using CsCl-ethidium
bromide gradient (Sambrook & Russell 2001). The microsatellite library enriched for
dinucleotide (CT) repeats was constructed following Lee et al. (2004). For those loci showing
multiple alleles, low stutter and robustness of interpretation, forward primers labelled with
6-FAM, HEX, or NED fluorescent dyes were synthesized and further used to confirm the polymorphic
loci using 24 large trees of S. lumutensis from Sungai Pinang.
Sample Collection and DNA Extraction
During the mapping process at the 8-ha study plot in Sungai Pinang, leaf or inner bark samples
were collected for individuals >1 cm dbh for genetic studies. In addition, a total of 40–48
representative samples were also collected from Pangkor Selatan, Lumut, Segari Melinting
and Teluk Muroh, using the transect-line sampling method, as explained by Lee et al. (2000).
The 54 individuals of S. lumutensis >20 cm dbh in Sungai Pinang were also used together with
the four half-sib families (B004, B005, B026, and B385) collected within the 8-ha study plot
for mating system and gene flow studies. Genomic DNA was extracted using the procedure
described by Murray & Thompson (1980), with modification.
Microsatellite Analysis
The samples were genotyped for four native microsatellite loci (Slu057, Slu110, Slu124 and
Slu175) and four microsatellite loci developed for S. leprosula (Sle111a, Sle118, Sle267 and
Sle303a; Lee et al. 2004). PCR amplifications and fragment analysis were performed according
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