58
N. Murakami et al.
was shown by the more comprehensive molecular work as a whole for Aspleniaceae
(Murakami 1995). A bootstrap analysis with 10 000 replications was performed in
order to estimate the reliability for various clades.
2.4 Ecological Observations
One leaf each was collected from 30 individuals of A. nidus along the loop trails
near Chikaniki Station, Mt. Halimun National Park, using a 6.5 m long fishing
pole with a small sickle on top. For each individual collected, their altitude, position on the tree (height from the ground), and degree of sun shade were recorded.
After returning to our laboratory at Kyoto University, a small portion of each leaf
was used for DNA extraction to determine their rbeL types. About 500 bp sequences
of rbeL were examined to identify the three types. Based on the recorded ecological
information and the obtained molecular data, the ecological preference of each
rbeL type was inferred.
3 Results
3.1 Variations in rbcL Sequence
We determined 1194 bp nucleotide sequences of l'beL. Three types of rbeL sequences were found from the 25 plants of A. nidus from Halimun. We called them
type A , Band C (Fig. 2). Among these three types, 30 sites of the rbeL sequences
varied.
We also determined the rbeL nucleotide sequences of the plants from Mt. Gede
and Bogor Botanical Garden. The type C was identified from the middle elevations
(1400-1900 m alt.) of Mt. Gede, but the plants from its higher elevations (19002400 malt.) and Bogor Botanical Garden (300 malt.) had very different rbeL from
any of the three types from Mt. Halimun.
3.2 Molecular Phylogenetic Analyses
The two equally most parsimonious trees (length = 146 steps; Consistency Index =
0.801; Retention Index = 0.851) were obtained by the Wagner parsimony method,
and their strict consensus tree is shown in Fig. 3. These three types were found to be
not closely related, when we included the plants of sect. Thamnopteris from the
oth.:r localities of the Old World. The type A is closely related to A. nidus from
Yunnan, China, and central Vietnam. The type C belongs to the same clade as type
A, but it was the most basally diverged from the other members. The type B made a
clade with A. australasieum senSll Holttum (1974) from Australia and New
Caledonia. We also analyzed the plants identified as A. nidus from Mt. Gede and
N. Murakami et al.
was shown by the more comprehensive molecular work as a whole for Aspleniaceae
(Murakami 1995). A bootstrap analysis with 10 000 replications was performed in
order to estimate the reliability for various clades.
2.4 Ecological Observations
One leaf each was collected from 30 individuals of A. nidus along the loop trails
near Chikaniki Station, Mt. Halimun National Park, using a 6.5 m long fishing
pole with a small sickle on top. For each individual collected, their altitude, position on the tree (height from the ground), and degree of sun shade were recorded.
After returning to our laboratory at Kyoto University, a small portion of each leaf
was used for DNA extraction to determine their rbeL types. About 500 bp sequences
of rbeL were examined to identify the three types. Based on the recorded ecological
information and the obtained molecular data, the ecological preference of each
rbeL type was inferred.
3 Results
3.1 Variations in rbcL Sequence
We determined 1194 bp nucleotide sequences of l'beL. Three types of rbeL sequences were found from the 25 plants of A. nidus from Halimun. We called them
type A , Band C (Fig. 2). Among these three types, 30 sites of the rbeL sequences
varied.
We also determined the rbeL nucleotide sequences of the plants from Mt. Gede
and Bogor Botanical Garden. The type C was identified from the middle elevations
(1400-1900 m alt.) of Mt. Gede, but the plants from its higher elevations (19002400 malt.) and Bogor Botanical Garden (300 malt.) had very different rbeL from
any of the three types from Mt. Halimun.
3.2 Molecular Phylogenetic Analyses
The two equally most parsimonious trees (length = 146 steps; Consistency Index =
0.801; Retention Index = 0.851) were obtained by the Wagner parsimony method,
and their strict consensus tree is shown in Fig. 3. These three types were found to be
not closely related, when we included the plants of sect. Thamnopteris from the
oth.:r localities of the Old World. The type A is closely related to A. nidus from
Yunnan, China, and central Vietnam. The type C belongs to the same clade as type
A, but it was the most basally diverged from the other members. The type B made a
clade with A. australasieum senSll Holttum (1974) from Australia and New
Caledonia. We also analyzed the plants identified as A. nidus from Mt. Gede and