Sex Determination in Calamus manan
 Choong Chee Yen, Wickneswari Ratnam, Kalaivani Nadarajah and Raja Barizan Raja Sulaiman


Rattan

Rattans are climbing palms belonging to the family of Palmae or Arecaceae and subfamily of Calamoideae. The word “rattan” is derived from the Malay “rotan”, the common name for climbing palm. There are about 600 rattan species of which can be grouped into 13 genera namely Calamus, Calospatha, Ceratolobus, Daemonorops, Eremospatha, Korthalsia, Laccosperma, Myrialepis, Oncocalamus, Plectocomia, Plectocomiopsis, Pogonotium and Retispatha (Dransfield et al . 1992). Among these genera, Calamus is the largest genus by number of species (Table 1). Of these many Calamus species, Calamus manan (Figure 1) is particularly commercially important for its valuable large-diameter cane used in furniture and handicrafts. Calamus manan or known locally as rotan manau is large-diameter rattan species with a stem (without sheaths) reaching 8 cm in diameter.


Table 1: The rattan genera

Genus

Number of species

Geographic distribution

Calamus

~370-400

Tropical Africa, India, Sri Lanka, China, Malesia, eastern Australia

Daemonorops

~115

India and China to western New Guinea

Korthalsia

~26

Indochina to New Guinea

Plectocomia

~16

Himalayas and south China to Malesia

Eremospatha

10

Tropical Africa

Ceratolobus

6

Peninsular Malaysia, Sumatra, Borneo

Laccosperma

5

Tropical Africa

Plectocomiopsis

~5

Lao, Thailand, Peninsular Malaysia, Sumatra, Borneo

Oncocalamus

4

Tropical Africa

Pogonotium

3

Peninsular Malaysia, Borneo

Calospatha

1

Peninsular Malaysia

Myrialepis

1

Indochina, Thailand, Myanmar, Peninsular Malaysia, Sumatra

Retispatha

1

Borneo
Source: Uhl and Dransfield (1987)


Figure 1: Massive plant of Calamus manan

Rattan manau has a wide distribution in Southeast Asia. It grows well on slopes of hill dipterocarp forest at altitude of 600-1000 m in Perak, Selangor, Kelantan,, Pahang, Terengganu, Negeri Sembilan, Sumatra and southern Thailand (Dransfield 1979). However, the over exploitation of the species for cane has limited its range in the forest. Calamus manan has solitary stem which can reach a length of 100 m. Cirrus located at the end of leaf is a climbing organ for the plant. The 3m-long cirrus covered with sharp and strong spikes is used to hook the neighbouring big trees for the support to climb up to the canopy for sunlight. The fruit of rattan manau is large compared to other rattan species with a size of 2-3 cm in diameter and is covered by reflexed scales.

Sex Identification

As most of other rattan species, rattan manau is a dioecious plant. The male and female inflorescences are found on different plants. Basically the morphologies of male and female plants are alike except their inflorescences. Female inflorescence contains fertile female flowers but sterile male flowers whereas the male inflorescence has only male flowers. The branching patterns of the male and female inflorescences are obviously distinct (Dransfield et al. 1992). The second order of rachilla in the female inflorescence bear flowers while the third order of rachilla in the male inflorescence bear flowers (Figure 2). This branching pattern in inflorescences is readily observed using a pair of binoculars for sex identification of rattan manau in the field.
Figure 2: Branching patterns of female (left) and male (right) inflorescences.

Rattan manau has a chromosome number of 2n=26. There is no indication of presence of sex chromosome in rattan manau. The sexes of rattan manau can only be determined when the plants start flowering. The rattan plants normally take 5-6 years of growing before first inflorescence appears (Wan Razali et al. 1992). Five to seven inflorescences per plant appear in one flowering season.


Importance of Sex Determination

Over exploitation on rattan manau resource from the wild has reached its critical stage. Our forests are no longer sustainable in supplying enough canes for rattan furniture industry. Therefore, cultivation of rattan in plantation seems to be a good way out for the problem. This may spare the wild rattan populations from being threatened. However, the identification of sexes in rattan manau can only be done when the plants start flowering at the age of 5-6 years. This seems to be too long for the establishment of nursery for supplying planting materials for mass cultivation of rattan manau in plantation. Optimum ratio of male and female plants in nursery is vital to maximize the generation of planting materials from seeds. Therefore, the application of molecular markers, particularly DNA markers, for sex determination in early stage is very much desired.


Molecular Approach

To achieve the sex determination at early stage of growing, molecular approach through the construction of cDNA library derived from floral tissues is attempted. It is expected that the floral mRNA of male inflorescence may differ to that of the female inflorescence of which may be used in determining the sexes of rattan manau. Rattan manau plants used in the study are located at F41, Bukit Lagong, Forest Research Institute Malaysia, Kepong. The rattan plants at F41 were planted in 1978. Most of the plants have reached a height of 10-15m. Therefore, scaffoldings were erected next to the selected plants for the purposes of close phenological observation and sampling of floral tissues for cDNA library construction. Floral tissues of different developmental stages of male and female plants were collected for total RNA extraction (Figure 3) and cDNA library construction. Eight cDNA libraries (4 male and 4 female) from different developmental stages of inflorescences were constructed. The size of the libraries ranged from 1100 to 18000 clones. The inserts of the clones were examined by means of direct PCR amplification from bacterial colonies. The insert sizes of the analyzed cDNA clones ranged between 500 to 2500 bp with an average insert size of approximately 1000 bp (Figure 4).
Figure 3: Total RNA extracted from floral tissues


Figure 4: Insert sizes of the cDNA amplified from colonies


Future Plans

The screening of the cDNA libraries are being carried out which involves DNA sequencing on clones, blasting and identification of EST sequences using GenBank database. We hope that DNA markers specific to sexes in rattan manau could be identified. These DNA markers will be subjected to testing on male and female plants before fully implemented.


References

Dransfield, J. 1979. A manual of the rattans of the Malay Peninsula. Malayan Forest Records No. 29. FRIM. Kuala Lumpur.

Dransfield, J., Wan Razali, W.M. and Manokaran, N. 1992. A guide to the cultivation of rattan. Malayan Forest Record No. 35. FRIM. Kuala Lumpur. pp. 39-46.

Uhl, N. and Dransfield, J. 1987. Genera Palmarum. Lawrence, Kansas, USA, Allen Press.

Wan Razali, W.M., Dransfield, J. and Manokaran, N. 1992. A guide to the cultivation of rattan. Malayan Forest Record No. 35.