Training Manualon Plant Taxonomy(Dicots)
in Southeast Asia

Training Manualon Plant Taxonomy(Dicots)
in Southeast Asia

Contributors
Dr. Dedy Darnaedi, Research Center for Biology, LIPI
Dr. Teguh Triono, Research Center for Biology, LIPI
Dr. Rugayah, Research Center for Biology, LIPI
Dr. Nanda Utami, Research Center for Biology, LIPI
Dr. Hary Wiriadinata, Research Center for Biology, LIPI
Dr. Marlina, Research Center for Biology, LIPI
Dr. Eiji Suzuki, Kagoshima University, Japan
Dr. Shiji Fujii, University of Human Environments, Japan
Dr. Edwino S. Fernando, University of the Philippines at Los Banos

Technical Editors
Dr. Dedy Darnaedi
Dr. Teguh Triono
Dr. Filiberto Pollisco, Jr.
Ms. Corazon De Jesus

EditorMr. Rolando A. Inciong

This TRAINING MANUAL ON PLANT TAXONOMY (Dicots)
IN SOUTHEAST ASIA is published by the ASEAN Centre for
Biodiversity though the Taxonomic Capacity Building and Governancefor Conservation and Sustainable Use of Biodiversity Project funded by
the Japan-ASEAN Integration Fund.December 2011

TABLE OF CONTENTS

Foreword .......................................................................................................................................................................................................................................................iv
Background and Rationale ...................................................................................................................................................................................................... v
Part I – The Training Course............................................................................................................................................................................................... 1
Objectives of the Training .............................................................................................................................................................................................. 1
Organization and Participants .................................................................................................................................................................................. 1
The Training Course Programme......................................................................................................................................................................... 2
Abstracts of Presentations .............................................................................................................................................................................................. 3
Training Outputs – Group Reports ................................................................................................................................................................ 14
Evaluation.......................................................................................................................................................................................................................................... 14
Part II – The Training Module ....................................................................................................................................................................................... 17
Methodologies for Plant Herbarium Collection ............................................................................................................................ 17
Cytology............................................................................................................................................................................................................................................... 20
Molecular Sytematics Laboratory Work ................................................................................................................................................... 25
Appendix...................................................................................................................................................................................................................................................... 41
Directory of Participants................................................................................................................................................................................................ 41
Training Course Programme ................................................................................................................................................................................... 45

Training Manual on Plants Taxonomy (Dicots) in Southeast Asia

iii

FOREWORD*I

t is a distinct pleasure for me to join you in the opening ceremony of the TrainingWorkshop on Taxonomy of Terrestrial Plants. This is a great event for the future ofyoung taxonomists in East and Southeast Asia, providing a venue to exchange knowledge,views and technical expertise in studying, evaluating and maintaining plant resources ofSoutheast Asia. It is common knowledge that floristically, our region is one of the richest
areas of the world, containing many native species of plants that are still untapped.
On behalf of the Indonesian Institute of Sciences (LIPI), allow me to welcome you all to
the Herbarium Bogoriense – the Research Center for Biology in Cibinong Science CenterLIPI. A special warm welcome is extended to the participants, observers and lecturers.
Indeed, it is a great pleasure for me to be here with you to witness this very important
course.
I really appreciate that this training is receiving enormous support from many parties
as reflected by the presence of distinguished delegates from the region and international
institutions.
We realize that the lack of trained human resources and inadequate capacities intaxonomy has been stressed as one of the obstacles in the implementation of Convention on
Biological Diversity commitments, especially in the ASEAN region. It is the mandate of LIPI
as a National Focal Point of the Subsidiary Body on Scientific, Technical and Technological
Advice (SBSTTA) and National Focal Point of the Global Taxonomy Initiative (GTI) andGlobal Strategy for Plant Conservation (GSPC) to promote basic and strategic research on
biodiversity especially in plant taxonomy.
I am very confident that this training-workshop will be an important milestone toenhance our expertise in the region, as well as strengthen our networking for the fruitfulcooperation among ASEAN taxonomists and East Asian experts.
At this juncture, I wish to stress that the training of young experts will give them
opportunities to hone their skills and gain additional knowledge in their field of expertise.
As for research,the participants will be given the chance to perform research work intaxonomy.
Finally, on behalf of the Indonesian Institute of Sciences, I would like to acknowledge
the generous support from the Ministry of Environment of Japan, ASEAN Center for
Biodiversity and Japan Wildlife Research Center. I would like also to express my gratitude
to Herbarium Bogoriense-Research Center for Biology-LIPI who organized and hosted thiscourse.
My heartiest gratitude is extended to all lecturers from the Philippines, Japan and
Herbarium Bogoriense, RC Biology-LIPI. I hope that the participants, lecturers, organizers
and sponsors will continue to communicate and strengthen your collaboration to increase
our understanding of our biodiversity.
*Welcome Remarks delivered by Prof. Dr. Ir. Bambang Prasetya, Deputy of Life Sciences, LIPI at the opening
session of the Training Workshop on Taxonomy of Terrestrial Plants, 16 February 2011iv

Training Manual on Plants Taxonomy (Dicots) in Southeast Asia

BACKGROUNDT

he lack of trained human resources and inadequate capacities in taxonomy has beenstressed as one of the obstacles in the implementation of Convention on Biological
Diversity (CBD)requirements, especially in the Association of Southeast Asian Nations(ASEAN) region. The dynamic growth of the ASEAN region and its neighboring countries
Japan, China and Korea (also referred to as ASEAN + 3) in recent years has increased
the pressure on their natural resources. Human activities, the driving force behind
the regional growth, threaten the biological resources. Lack of scientific information
on biodiversity in this region is a crucial issue in the assessment and prediction of
biodiversity changes, caused mainly by the lack of taxonomic capacity in data collection
and analysis.
During the Ninth Meeting of the Conference of the Parties to the CBD (CBD-COP9), theProgramme of Work on the Global Taxonomy Initiative (GTI)with deliverable outcomes
was adopted (Decision IX/22). The Programme of Work, as revised, was adopted in CBD
COP10 through Decision X/39.Parties to the Convention were urged to promote/carryout the Programme of Work for the GTI through coordination of its implementation withexisting national, regional, sub-regional, and global initiatives, partnerships and institutions;designation of national GTI focal points; provision of updated information about legal
requirements for exchange of genetic/biological specimens; about current legislation andrules for access and benefit-sharing in terms of the needs for the GTI; and initiatives ofsetting-up of national and regional networks to aid the Parties in their taxonomic needs in
implementing the CBD.
Adequate taxonomy is one of the necessary fundamental tools required for the
global community to be able to implement the Millennium Development Goals and thedevelopment targets from the World Summit for Sustainable Development. Without
adequate long-term investment in the human, infrastructural (including, importantbiological collections) and information resources necessary to underpin the science oftaxonomy, the now well-recognized taxonomic impediment will continue to prevent
adequate implementation of sound, scientifically-based sustainable, environmental
management and development policies.
It is in this context that the project “Taxonomic Capacity Building and Governance forConservation and Sustainable Use of Biodiversity” was approved by the Japan-ASEAN
Integration Fund in July 2010. An Inception Meeting/Workshop was conducted in Manila,
Philippines on 31 August – 01 September 2010 to provide direction and work out the detailsof the project.The workshop identified three training topics to be conducted, one of which is
the taxonomy of plants.
The main objective of this manual is to provide a clear, simple and concise technicalhandbook for young researchers in plant taxonomy. The handbook aims to increasescientific expertise, and improve skills in management of specimens, writing, and in
preparing taxonomic publications. This manual is an attempt to bring together the current
knowledge on the theory and practice on plant taxonomy, from morphology, anatomy,
geography, ecology, cytology and molecular biology.

Training Manual on Plants Taxonomy (Dicots) in Southeast Asiav

Many people have been involved in the production of this manual, the content of
whichwas based on the training-workshop conducted at the Herbarium Bogoriense, RC
Biology-LIPI, and the experiences of the Herbarium Bogoriense staff. It is hoped that the
manual will become a widely used and accepted handbook to help in-services training inmany institutions of the different ASEAN member States.
The organization of this manual is in two parts. Part I provides a full report of theTraining Course conducted at the Herbarium Bogoriense, RC Biology – LIPI in Bogor,
Indonesia, from 16 to 23 February 2011. Part II is a Training Manual largely based on the
training course.
We are indebted to the Japan ASEAN Integration Fund for funding this manual andto the ASEAN Centre for Biodiversity (ACB) and the Research Center for Biology-LIPI fororganizing the course. Our gratitude goes to the following staff for their contributions:Dr. Teguh Triono, Dr. Rugayah, Dr. Nanda Utami, Dr. Hary Wiriadinata and Dr.Marlina
of Herbarium Bogoriense, RC Biology-LIPI; Dr. Eiji Suzuki from the Graduate School of
Science and Engineering, Kagoshima Universityand Dr. Shiji Fujii fromtheUniversity of
Human Environments inJapan; and Dr. Edwino Fernando of the College of Forestry andNatural Resources at the University of the Philippines Los Banos for their lecture materials
during the Training-Workshop.

vi

Training Manual on Plants Taxonomy (Dicots) in Southeast Asia

PART I – THE TRAINING COURSE

Objectives of the Training Workshop
Generally, the training workshop aimed tocapacitate participants in the rigors oftaxonomy especially on the dicotyledonous plants group. Specifically, the training
workshop:
1. Introduced the participants to the taxonomy of terrestrial plants particularly on
selected families of dicots
2. Familiarized the participants with the general biology of these plants
3. Upgraded the taxonomic skills of the participants on the following:a. Methods of morphological observationb. Sample collection, processing and managing
c. Databasing
4. Introduced the participants to advanced taxonomic methodologies such as moleculartechniques, bar coding and databasing
5. Provided hands-on experience through a field exercise in collection, DNA coding andmolecular analysis
The outputs of the training workshop were the following:
1. Skills of the participants in terrestrial plants taxonomy upgraded
2. Advanced taxonomic methodologies introduced
3. Hands-on experience in collections management, cataloguing and storage provided

Organization and Participation of the Training Course
The training
course wasorganized by the
ASEAN Centre
for Biodiversity(ACB) and theMinistry of theEnvironment, Japan(MoE-J), and hosted
by Herbarium
Bogoriense,
R.C.Biology, CSCLIPI in Bogor,
Indonesia.
The course requiredparticipants from each ASEAN Member State who had backgroundin botany, plant ecology and related fields, not older than 35 years old, and with academicor government position working on terrestrial botany or plant ecology.

Training Manual on Plants Taxonomy (Dicots) in Southeast Asia

1

There were 15 participants representing eight ASEAN Member States, plus three
additional participants from Indonesia andan observer from China.
There was a balance in the number of male and female participants. The age distribution
was also very much within the range of the age requirement, except for two participants
who were above the 35 years old age limit. Professionally, nine participants were working
with academe or research institutions, and tenwith the government. The directory ofparticipants can be found in Appendix 1.

The Training Course Programme
The training course on plant taxonomy had three methodologies: lectures, specimen
management, and hands-on field collection. A copy of the course programme can be foundin Appendix 2.
The lectures were on the following:
1. General lectures on:a. Taxonomy
b. Morphology
c. Biogeographyd. Cytology
e. Bar coding
f. Molecular analysis
g. Specimen collection methodologies
2. Special lectures on the following dicot families:a. Fabaceae / Legumes
b. Rubiaceae
c. Rosaceae
d. Ericaceae
e. Sapotaceae
f. Euphorbiaceae
g. Cucurbitaceae
h. Myristicaceae
The participants were divided into
groups to work on cytology and on
DNA extraction as part of the specimen
management. For the field collection
exercise, a field trip to the Cibodas
Botanical Garden was conducted.
As a culminating activity, theparticipants chose specific dicot families
for their group presentations and reports.
Putting together their learning from thelectures, they collected samples from thebotanical garden, identified the specimen,
made the taxonomical keys based onmorphology, and provided the plant
description.
A CD was distributed to the participants containing all the presentations of the groups,
the lecturers, as well as the pictures taken throughout the training course. In addition, theDarwin Core, a biodiversity information database standard to facilitate exchange about
geographic occurrence of species, and integrating this with other information about theorganisms was presented and given to the participants.

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Training Manual on Plants Taxonomy (Dicots) in Southeast Asia

Abstracts of Presentations
Plant taxonomy is a basic and fundamental research in understanding plant diversity.
A specific technical course is usually needed for each group of plants, such as courses on
Monocotyledoneae, Dicotyledoneae, Ferns, Bryophyte and Fungi. The broad scope of thecourse, from morphology to molecular analyses, requires sufficient period of time for its
conduct. For a two-week course, a special topic can be selected as a main course, while theother topics can read to develop general understanding. For anyone interested in molecularanalyses, he/she can directly follow the technical manual for DNA analyses; however,additional subjects are required to understand the relationship among the other subjects.
Genetic information through cytology and molecular analyses are interesting andexciting subjects for young scientists. However, in any taxonomic work, one needs tounderstand other subjects such as morphology, anatomy, paleontology, ecology andgeography etc. During the training course, all subjects were discussed.
The training course was divided into lectures (basic taxonomy, morphology, dicot
families), laboratory work (herbarium work, cytology, molecular analyses), database,and field collection. The complete lectures during the training course are provided as
appendices (Appendix 1).

Plant Taxonomy (Introduction)
Dr. TeguhTriono, Head of Plant Taxonomy Group, RCBiology-LIPI, Indonesia
The foundation principles of taxonomy stem from the natural need for classification,
recognizing that a ‘name is a key to information’.
“The urge to classify is a fundamental human instinct; like the predisposition sin, it
accompanies us into the world at birth and stays with us to the end.” (A.Tindell Hopwood
1959 in Davis & Heywood 1963).
The aims of taxonomy are for identification and communication; to provide naturalclassification; and to reconstruct evolutionary pathway. The phases in taxonomy
are exploratory phase (identification); consolidation phase (adding more materials);
biosystematic phase (taxonomic evidence); and encyclopaedic phase (drawing evolutionary
relationship).
The common terminologies being used in taxonomy are identification, description,
classification and key, and phylogeny. In plant classification, relationship is being assessed,delimiting taxa and assigning the rank of a species. Criteria for classifying plants would
include genetically fixed variation; based on correlation and discontinuity of characters;
consideration of all materials used; covers an entire geographical range; and having
consistent treatment.
Characters as used in taxonomy may be analytic and synthetic, qualitative andquantitative, good and bad. Taxonomic evidences would also need to be produced, sinceclassification will often be based on total evidence, such as morphology, cytology andanatomy, metabolites and DNA, and field, herbarium and library specimens. Taxonomic
products contain information, and these could be in the form of a checklist, notes, flora,monograph or a phylogenetic paper.

Plant morphology of Dicotyledonae
Dr. TeguhTriono, Head of Plant Taxonomy Group, RCBiology-LIPI, Indonesia
Plant morphology simply refers to “the way in which plants are constructed”. This isnecessary for quick and easy identification, in writing descriptions, in using literature (keys,
plant descriptions), laying-out, collecting, and for field notes. Careful observation is always
the key. A set of standard works is usually adopted, such as that of Hickey (1979) toTraining Manual on Plants Taxonomy (Dicots) in Southeast Asia

3follow leaf morphology, and the Systematics Association Committee (1962) for shapes. Key
characters of plants, in morphological units, include habit, sap, leaves, stipules, hairs andglands, flowers, and fruits.

Biogeography
Dr. Edwino S. Fernando, Professor, University of the Philippines - Los Banos,
Department of Forest Biological Sciences, UPLB, Philippines
Plant geography or phytogeography is often regarded as one of the oldest of plantstudies. It deals with the distribution of plants over the Earth’s surface, past and present,
and is the science that attempts to describe and interpret the geographic distributions oforganisms. A common concern in plant biogeography is the distribution and dynamics ofplant biodiversity, often simplified in terms of numbers, or proportions of endemic species.
The Earth’s surface is often divided into geographical units based on similarities anddifferences in the occurrences of species and ecosystems such as Biogeographical realm,
Biogeographical provinces, Biome, Ecoregion, Vegetation, Flora, Floristic areas, andEndemism, among others. A general question in historical biogeography is why not a
uniform Flora worldwide? To which the obvious reply would be the differences in biotic
and abiotic environmental factors in various parts of the world.
Disjunctions – which are natural gaps in the geographic ranges of taxa – accountfor the many reasons why in seemingly identical conditions of different areas there arestill different floras, as well as for the discontinuous distributions of species. Causes of
disjunction may be chance dispersal and vicariance. Phylogenetic analyses may reveal theevolutionary relationships among the resultant lineages and allow for reconstruction of theevents that resulted in their original separation from one another.
Phylogeography is the study of the geographical distribution of genealogogical lineages
as determined by DNA sequence variation from individuals across a species range.
Conservation biogeography is the application of biogeographical principles, theories andanalyses, being those concerned with the distributional dynamics of taxa individually andcollectively, to problems concerning the conservation of biodiversity.

Plant age
Dr. Eiji Suzuki, Professor, Earth and Environmental Sciences, Graduate School of Scienceand Engineering, Kagoshima University, Kagoshima, Japan
Plants live longer than animals. The oldest recorded plant, the Pinus aristata in theUSA, was recorded to be 4,862 years old, while the longest living animal, the turtle, wasrecorded at 152 years old. In a cold climate zone, many trees have annual rings, and these
are counted to estimate the age. The ages of trees without annual rings are estimated by C
methods. Age of animals is mostly from breeding records in zoos. For whales, their earwax
accumulates, making annual scars, which are then estimated as records of their age. Shelled
species, on the other hand, make annual scars in their shells.
Some animal physiologists, however, contest that plants do not live longer than animals.
Plants, they say, live for only a few decades because, for instance in tree cells, the vessels ortracheid are dead cells from the beginning, and that the living cells, such as those in leaves
and phloem, live for only 1-30 years. In animals, however, most of their cells are alive,with the same cells living throughout their lifespan. The brain cells of a 150 year-old turtle
is actually alive throughout its life span. The difference of life between plants and animals
are in (1) structure of cells; and in (2) reproduction. In determining the difference between
angiosperms and gymnosperms, the age, physiology and evolution should be considered.

4

Training Manual on Plants Taxonomy (Dicots) in Southeast Asia

Cytology
Dr. DedyDarnaedi, Plant Taxonomist, Senior Staff, Herbarium Bogoriense,
RCBiology-LIPI, Indonesia
Plant systematics serve as a major unifying force in biology because it uses datafrom a multitude of discipline, such as molecular biology, biochemistry, comparative
phytochemistry, electron microscopy as well as morphology, ecology, biography to develop
a framework upon which classification can be developed. Indeed without the system
of nomenclature provided by plant taxonomy, there could be no comparative referencein naming of plants. Cytology is more commonly known as cell biology, and is also
an important discipline relevant to taxonomy. Specifically, the genetic or chromosome
information in the cell, such as the number of chromosomes in each cell, the size ofchromosome, centromere position, and the behavior of chromosomes, are being studied.

Molecular systematics
Dr. Marlina, RCBiology-LIPI, Indonesia
There are several ways to collect material for DNA extraction. The ultimate thing toremember is how we prevent the cells from undergoing destruction. Collecting methods
may be through Silica gel, FTA PlantSaver Card (Whatman), Liquid Nitrogen and CTAB
Solution. The polymerase chain reaction (PCR) is a technique widely used in molecularbiology. As PCR progresses, the DNA generated is used as a template for replication. This
sets in motion a chain reaction in which the DNA template is exponentially amplified.

Database IBIS
Mr. Bambang Hartoko and Mr. Basuki Bundi Mulyono
The Indonesian Biodiversity Information System (IBIS) is an application developed by
the Center for Research Biology-LIPI. Developed by using open source software, the IBIS isused to manage data of specimens of flora, fauna and microorganisms (specimen data arederived from flora specimens, fauna specimens, and living micro-organisms specimens). Its
development began in 1991.The migration of data and records took place from 1994 to 2003,
and since then the IBIS has been utilized and implemented. The last version of IBIS wasdeveloped in 2011, consisting of five tables – Collection, Specimen, Locality, Determination,
and Component.

Training Manual on Plants Taxonomy (Dicots) in Southeast Asia

5THE DICOT FAMILY
Ericaceae
Dr. Edwino Fernando

Ericaceae are mostly trees and shrubby plants, rarely lianas or herbs, sometimes
epiphytic, mycorrhizal, some achlorophyllous. Leaves are simple, often spiral(including pseudoverticillate), decussate or whorled, and always exstipulate, with glands
below, venation pinnate (sometimes with three or more veins from the base). Flowers inbracteate racemes, sometimes solitary, often pendulous, usually actinomorphic, usuallybisexual; petals connate and typically campanulate, funnel-shaped, cylindrical, tubularor urceolate; stamens are five to ten, sometimes free or adnate to the corolla, the anthers
often dehiscing by apical pores and sometimes have appendages; ovary five-locular.
Fruit is capsule (loculicidal or septicidal), berry, or drupe; seeds are numerous, usuallysmall, sometimes winged. Many species are edible, as well as used as ornamental
plants. It usually grow in elevations of 1,500 meters to very high elevation such as alpine.
They usually grow as shrub, or small trees, but in the lower elevation they can grow as
epiphytes. They have very attractive, colorful flowers,have potential as an ornamental plant,
and have medicinal properties.
Spot character:• Always: Leaves spiral (incl. pseudoverticillate), exstipulate; flowers hermaphroditic;
corolla tubular or urceolate, anthers opening by apical pores, style 1; seeds numerous.
• Usually/often: Woody; leaves glandular below, venation pinnate, sometimes with 3 ormore secondary veins from base; flowers actinomorphic; ovary 5-locular; stamens 10,
anthers with appendages.
• Striking features: Saprophytes (Andresia, Monotropastrum); leaves lepidote (mostRhododendron); glands on leaf margin at base• Different from: Epacridaceae: venation parallel.
One group who choses Ericaceae studied some species of Rhododendron. In Java, there
are 13 species. Near Cibodas Botanic Gardens the wild Rhododendron usually grows as
epiphyte on the branches of big trees, several species can be found in the gardens as
terrestrial shrubs.

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Training Manual on Plants Taxonomy (Dicots) in Southeast Asia

Sapotaceae
Dr. Teguh Triono

Identifying the plant morphology of Sapotaceae includes determining the presence orabsence of cuts or wounds and color of the sap or exudate. Leaves could be simple,
opposite/alternate, crowded/dispersed, and with margins of the entire plant.
Sapotaceae may be hermaphrodite or unisexual, with the ovary of its flowers either
superior (hypogynous) or semi-inferior (epigynous).

Begoniaceae
Dr. HaryWiriadinata

Begoniaceae is a big family containing more than 1,500 species found wild throughout
tropical and subtropical Asia, Africa and America. It is characterized by its herbaceous habit,
rarely woody stem, asymmetric leaf, unisex flower and many tiny seeds. Many species havebeautiful leaves so they become ornamental plants which have commercial importance in thehorticultural trade. Over 10,000 kinds of hybrids and cultivars are available.

Training Manual on Plants Taxonomy (Dicots) in Southeast Asia

7• Herbaceous plants, sometimes tuberous, monoecious.
• Leaves simple alternate.
• Stipules always present on the base of the petiole, sometimes caducous.
• Inflorescence basically cymose or racemose.
• Flowers always uni-sexual; the sepals, petals sometimes similar. In male flowers
usually have 2 outer sepals and 2 smaller inside sepals, free; stamens numerous,
filament free, anthers are spherical or hemispherical cluster, dehiscing longitudinally.
The female flowers usually have 4-5 sepals; ovary inferior which has 3 locules(chambers); ovules many per locule. Fruits a capsule rarely berry.
Spot character• Always: Herbs, stem succulent, sometimes woody at base; leaves simple, alternate,
asymmetrical, stipulate; flowers unisexual (plants monoecious); stamen many; ovary
inferior, 3 locular, ovules numerous.
• Usually/often: Sepals and petals free, fruit 3-winged.
• Different from: The family is allied to the Datiscaceae (huge trees); sterile Begoniaceae
might be mistaken for Balsaminaceae (exstipulate, symmetric leaves) - Gesneriaceae(Cyrtandra): leaves opposite – Urticaceae: Cystoliths.
One group chose Begoniaceae for their study due to its potential as ornamental plant.
There are several species that occur in the wild, few are in botanic garden collections. Is
there a natural hybrid between Begonia robusta and B. multiangula since they grow in thesame ecological niche? The dense red hairs on the young leaf of Begonia robusta sometimes
loses an old leaf. Begonia isopteran is characterized by its equal 3-wing fruit.

FabaceaeDr. Hary Wiriadinata

Majority of the members of the Fabaceae family occurs from sea level to mountainous
flora.

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Training Manual on Plants Taxonomy (Dicots) in Southeast Asia

Rosaceae
Dr. Hary Wiriadinata

The majority members of Malesian Rosaceae belong to the mountain flora and occur
only above 1000 meters above sea level. It is a large family with worldwide distribution.
• Tree or herbaceous plants.
• Leaves always spirally arranged, sometimes distichous, simple or compound.
• Stipules on the twig or on the base of the petiole, free or adnate to petiole, rarely
absent. Inflorescence various.
• Flowers always bisexual and actinomorphic; hypanthium usually very distinct, fromsaucer-shaped to tubular or campanulate; the sepals petals and stamens inserted on its
rim, its inside usually lined by a nectariferous disk.
• Sepals usually 5, free, sometimes an epicalyx present.
• Petals usually 5, free, from large and showy to small and not or hardly distinct fromsepals or absent.
• Stamens usually numerous, filament free, anthers bilocular, dehiscing longitudinally• Pistil (s) one to numerous, free.
• Ovary (ies) superior to inferior; style present; ovule (s) one to several per locule.
Spot characters:• Always: Leaves spirally arranged, stipulate (except Spiraea); flowers actinomorph andstyle present.
• Usually: Leaves serrate, flowers hermaphrodite, stamens numerous.
• Striking features: leaves drying reddish brown, black glands below, fruit often weakly
bilobes (Prunus); hooked bristles on hypanthium (Acaena, Agrimonia); spiny plants(Rosa p.p., Rubus.)• Different from: Chrysobalanaceae: always trees, flowers more or less zygomorphic,style more or less exentric. Symplocaceae: exstipulate, ovary inferior; Saxifragaceae:
usually exstipulate.

Training Manual on Plants Taxonomy (Dicots) in Southeast Asia

9

During the field trip, one teamfound and studied some species of the genus Rubus.
This groupis characterized by spiny stem, compound leaflets, persistent calyx, 5 free white
petals, many stamen, and many pistils. Fruit are berries.

Rubiaceae
Dr. Hary Wiriadinata

The family Rubiaceae is trees, shrub, vines or herbaceous plants that are sometimes
ant-inhabited. Members of this family can be found from sea level to mountainous areas.
• Trees, Shrub, Vines or herbaceous plants, sometimes ant-inhabited (as Hydnophytum,
Myrmecodia, Myrmeconauclea).
• Leaves simple or compound, sometimes opposite arranged, sometimes distichous.
Stipules on the twig or on the base of the petiole, free or adnate to petiole, rarely
absent. Inflorescence various.
• Flowers always bisexual and actinomorphic; hypanthium usually very distinct, fromsaucer-shaped to tubular or campanulate; the sepals, petals and stamens inserted on
its rim, its inside usually lined by a nectariferous disk.
• Sepals usually 5, free, sometimes an epicalyx present.
• Petals usually 5, free, from large and showy to small and not or hardly distinct fromsepals or absent.
• Stamens usually numerous, filament free, anthers bilocular, dehiscing longitudinally.
• Pistil (s) one to numerous, free.
• Ovary (ies) superior to inferior• Style present.
• Ovule (s) one to several per locule.
Spot characters:• Always: Leaves decussate, including verticillate (sometimes one leaf of a pairreduced), simple, entire, interpetiolar (rarely intrapetiolar) fused stipules; corolla
tubular, stamens isomerous, alternate with lobes.
• Usually/often: Leaves with raphides; flowers hermaphrodite, actinomorphic, corolla 4
or 5 lobed; ovary inferior, 2-locular, placentation axile.

10

Training Manual on Plants Taxonomy (Dicots) in Southeast Asia• Striking features: Leaves of a pair unequal, flowers like those of Solanum(Argostemma); stipule ochreate, ovary superior (Gaertnera); climber with hooks(Uncaria); one sepal enlarged and showy (Mussaenda); epiphytes with swollen stems
inhabited by ants (Myrmecodia, Hydnophytum)• Different from: Caprifoliaceae: leaves exstipulate; Loganiaceae: no raphides, ovary
superior; Rhizophoraceae: petals free.
There are several genera and species along the trail to Cibeureum waterfall such as
Borreria, Lasianthus, Mussaenda, Mycetia, Pavetta, Psychotria, Urophyllum and Wendlandia. The
genus Lasianthus and Urophyllum have similar morphological characters and they have alsosimilar habitat. Lasianthus sometimes misidentified as Urophyllum. In this training somespecies of Lasianthus were investigated in more detail.

Cucurbitaceae
Dr. Rugayah

The family Cucurbitaceaehas a species diversity of about 900 species belonging to 120
genera, with 37 genera found in Malesia. It is distributed all over the world, predominantly
in tropical areas, with more than 90 percent of the species found in Africa, Madagascar,
Central and South America and Southeast Asia and Malesia. Its value is usually as edible
fruit and seed (Cucurbita, Cucumis, Citrulus, Benincasa, Sechium, Luffa, Lagenaria, Momordica,
Cyclanthera, Coccinea), and for medicinal purposes (Trichosanthes, Momordica) as well.
Cucurbitaceae is mostly found in primary or less disturbed forest, seeming to prefergrowing in shady places, wet condition, or openareas along the riverside. Some may beclimbing on other plants, creeping on theground or rocks. The plants are mostly unisexual,
monoecious or dioecious, so both male and female flower per plant have to be collected.
Collecting samples in the morning is better. Most species are nocturnal, meaning theiranthesis flowers take place at night till the morning of the following day. Wet collection
and taking photographs arestrongly recommended because the flowers are showy, but
thin and easily damaged.The fruit is also showy, varying in shape and color, juicy (somehave hardy pulp).

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Balsaminaceae
Dr. Nanda Utami

The Balsaminaceae is composed of two genera – the monotypic Hydrocera and theprolific Impatiens, which are commonly referred to as the balsams. The Hydroceraaresemi-aquatic plants, growing in swampy places, watersides and lakes. They canbe found insouthern India, Ceylon, and parts of Southeast Asia. Impatiens is agenus of about 850-1000
species of flowering plants. The genus is concentrated in mountainous regions of Southeast
Asia, south China, India and Africa. The scientificname Impatiens is Latin for impatient,in reference to their seed capsules. When thecapsules mature, they “explode” whentouched, sending seeds several meters away.This mechanism is also known as “explosive
dehiscence”.

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Training Manual on Plants Taxonomy (Dicots) in Southeast Asia

Fagaceae
Dr. Shiji Fujii

Distinguishing morphological characters of Fagaceae include the fruit and the cupule.
The cupule is a mysterious organ. Many hypotheses had been proposed to explain its
origin – intercalary growth, shoot axis, etc. One hypothesis says that it originated fromsterile shoots (lateral shoot axis). In its primitive form, there are three or more flowered
dichasial cyme, and with a lobed cupule (where each valve is related to a sterile shoot).
There are five hypothetical patterns of distributions of species along an environmental or
geographic gradient: (a) Species are distributed as discrete communities that replace eachother abruptly; (b)Species are not segregated into communities, but some sets replace eachother abruptly; (c) Species are distributed as discrete communities, which gradually replaceeach other; (d) Species behave as if they are independent of each other, neither associating
in discrete communities nor replacing each other abruptly; and (e) Most species are nested
within the ranges of a few dominant species, but otherwise occur independently of eachother.
References:
Backer, C.A. & RC Bakhuizen vd Brink Jr. 1963-68. Fl, Java (3 vols). Noord. Batavia.
Hughes, M. 2008. An Annotated Checklist of Southeast Asian Begonia. Royal Botanic Garden Edinburg.
164 pp.
Kalkman, C. 1993. ROSACEAE. Flora Malesianaser. I, vol 11 (2): 227-351.
Keng, H. 1978. Orders and Families of Malayan Seed Plants. Singapore Univ. Press. 437 pp.
Kiew, R. 2005. BEGONIAS of Peninsular Malaysia. Natural History Publication (Borneo).
308 pp.
Nielsen, I.C. & H.C. Fortune Hopkins. 1992. MIMOSACEAE (LEGUMINOSAE-MIMOSOIDEAE). Flora
Malesianaser. I, vol 11 (1): 1-226.
Polhill, R.M. &P.H.Raven.(ed.) 1978. Advances in Legume Systematics Part 1.Royal Botanic Gardens,
Kew. 425 pp. í-xxí.
Van Balgooy, M.M.J. 1998. Malesian Seed Plants. Vol.2 - Portraits of Tree Families.

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Training Outputs
The participants chose specific dicot families for their group presentations and reports.
Putting together their learning from the lectures, they did the collection from the botanicalgarden, identified the specimen, made the taxonomical keys based on morphology,
and provided the plant description. The participants worked on the families Ericaceae,
Begoniaceae, Rosaceae, and Rubiaceae. They produced a report on the families assigned tothem and made a presentation on the last day of the training. The reports of the groups arefound in Appendix 4.

EVALUATIONA pre- and post-training evaluation was conducted with the participants. A summary of
their answers is found below.

Pre-training Evaluation
What skills/knowledge/behavior do you want to develop by attending this training workshop?
While majority of the participants expressed a general expectation to gain more
knowledge on plant taxonomy, a few expressed their desire to develop specific skills on
identification, classification, morpohology, cytology, molecular analysis, curation, collection,
databasing, barcoding, herbarium work. Some mentioned that they wanted to develop
specific knowledge on seed plants, freshwater (plants), vegetables and tropical trees.
What do you expect to see/hear/feel differently by developing the above skills/knowledge/behavior?
The participants expressed that they expect to be more confident and efficient, which are
the skills needed tobe a professional taxonomist. Some participants said they expected theirinitial impressions about the difficulty of cytology work, molecular studies and other planttaxonomy work will be unfounded, and that their skills would actually improve. Other
participants expected that in interacting and sharing experiences and knowledge with theother participants, their social skills will improve.
How will it benefit your job performance by developing the above skills / knowledge / behavior?
Most of the participants are working in herbariums, botanical gardens or nationalparks in their countries. They all believed that the training will benefit their work andjob performance, as the new skills will support their tasks (research, identification andcollection of plants, curation and management). Some participants expected that by
improving their job performance, they will be able to speed up the completionof theirprojects, and may even attract potential funding agencies to support further research.
How do you want others to relate to you after attending the training workshop?
After attending the training workshop, the participants expected to have continuous
contact with their fellow participants, with some hoping for strong collaboration andregional networking. One simply wanted others to relate to him as a professionaltaxonomist/systematist.

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What do you feel you are currently not achieving due to the underdevelopment of the above skills/
knowledge/behavior?
Although most of the participants were positive in expressing that underdevelopmentof skills is not necessarily a problem, they believed they can still work and improve on
it.Some of them expressed that their learning and skills related to work are not enough,thus hindering them from completing projects on time. A participant said underdeveloped
skills havecaused low self-esteem.
What are your personal learning goals? What do you really want to learn from this training
workshop?
The participants had common personal learning goals – to learn skills related to work
andto develop/establish relationships with other institutions and people.The Cambodian
participant specifically mentioned “to protect and conserve forest plants in Cambodia forsustainable use”.
What do yoursupervisors expect from you after attending the training workshop?
While most of the participants expressed that their supervisors expect them to improveand enhance their skills in taxonomy, as well as learn new and advance taxonomic
methodologies, others said they were expected to train their colleaguesand share theirlearning from the training. They were also expected to network with other taxonomists andorganizations.
What other expectations do you have of this training workshop?
Other expectations raised by the participants included knowing the ‘science culture’ ofother participants; developing an international network not just for taxonomy but creating
friendships as well; and other activities that would support their existing knowledge onplant taxonomy, among others.

Post-training Evaluation
Which of the training lectures/sessions need to be improved? Please elaborate which aspect needs
to be given attention (time allotment, clarity of topic, instructor/trainer expertise, processes/
methodologies used).
The participants suggested that more time be allotted to lectures. They emphasized theusefulness of hands-on work (laboratory, field collection) combined with the lectures. There
were mixed review on the trainers, with participants appreciating the expertise of some of
the trainers, but suggesting that some of the trainers needed to improve their lecture on
certain plant families.
Kindly provide your recommendations to further improve the training lectures/sessions.
The participants recommended that more detailsshould be added to each topic, such as
on the principles of plant taxonomy, nomenclature, database and more in-depth lecturesand hands-on training on specific or focused families, instead of including many families ina brief period of time.They also recommended inviting more experts as trainers.

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Make a list of learning and results from each goal (as stated in the pre-evaluation form).
Most of the participants appreciated learning how to identify the dicot families, do keys
and descriptions, as well as learn and experienceabout cytology and molecular laboratory
work. Their expectations and goals to learn more about plant taxonomy were clearly met,
but the participants reiterated that there was very limited time to absorb and understandeverything.
What was your biggest learning experience from this training?
The answers ranged from a specific topic/lecture that the participants understood better(cytology, molecular systematic, plant identification, bar coding), the skills that they learned(field work collection, plant identification, laboratory work), and social skills they gained(networking, knowledge sharing, friends). One participant pointed out how he ‘learned somany things in a short time, and still able to make a report’.
What skills/knowledge/attitude do you have now, that you didn’t have before attending the training?
What are your plans after this workshop?
The participants enumerated a number of specific skills and knowledge that they haveacquired in attending the training, such as making a key, identifying the species in the dicot
family, doing cytology work and molecular analyses, among others. Attitude-wise, they said
they wereinspired to be better equipped with more plant knowledge, as the experience will
be wisely applied to their work. They also acknowledged that the training allowed them towork better within a group, helping them to be better listeners in order to work within ateam.
What changes will you do to apply what you have learnt from the training workshop into your dayto-day job?
There was a good mix of knowledge, skills and attitude that the participants learned
from the training, which they are eager to apply to their day-to-day jobs. Participants
expressed that they will try to identify the unidentified collections that they have in theirbotanical gardens, and expand identification to cellular/cytology level. One participantwas, on the other hand, eager to prepare and submit proposals to funding agencies fortaxonomic researches. Other changes that they will apply include being more criticalregarding plant taxonomy problems, and to contact plant taxonomists in creating/developing a network.
What support do you need from your supervisors and colleagues to realize the plans you mentioned
above?
The participants said they needed financial support (for their researches, especiallychemicals and equipment for their cytology studies), freedom (to conduct their researches,studies, as well as to attend more training and have more exposure), and support to theirideas, sharing of information, peer-to-peer review of studies, among others.
Do you have any other comments about the training workshop?
With a general sentiment that the training course was overall satisfactory, theparticipants emphasized the need for more time for the training course – for the lecturesand field work, in order for them to practice taxonomic skills and in studying a plant
species.

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Training Manual on Plants Taxonomy (Dicots) in Southeast Asia

PART II – THE TRAINING MODULE

T

his Training Module is divided into four parts. The first focuses on Methodologies for
Plant Herbarium Collection, the second part on Cytology, and the third section is an
introduction to Molecular Systematics and Laboratory Work. The last part of the module
provides brief morphological characters of selected dicotyledons.

Methodologies for Plant Herbarium Collection
The Malesia region (Malaysia, Singapore, Indonesia, Papua New Guinea, the Philippines
and Bismarck Islands) is a megadiverse region. About 250 to 300 species of flowering
plants, or 10 percent of higher plants, occur in the region. In the Herbarium Bogoriense,
there are about three million herbarium specimens and references about plant taxonomy.
Even if the herbarium specimens are stored alphabetically in new cabinets in comfortableair-conditioned rooms, it would take time to study them all within a short period. The
easiest method to study higher plants within a short period is by studying the living
selective group of genus with a small number of species, making herbarium specimens, andstudying their morphology.
First, write all characters and character state of single individual plant, from generalcharacter to specific character, from base to top. Do the same to second individual, third
individual, and so on for one species, and find out the constant characters. Compare thecharacters and character states of different species, and use those differences for your key toseparate the species.

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For the training course in Bogor, herbarium collections weremade from the field at the Cibodas Botanic Gardens, and along
the trail to Cibeureum waterfall within Mt. Gede Panggrango
National Park at elevation 1400-1600 m high altitude. Since too
many plants occur in those areas, the objective of collection was
limited to several relative species of selected families such as
Rosaceae, Ericaceae, Rubiaceae and Begoniaceae that grow wildin that area, with three or more species of one genus of thosefamilies. As introduction to every selected family of higher plants
in this manual, for each selected family make some notes of thedelimitation characters, spot the characters, and compare different
characters of one family with a close relative family. Since thefour large families contain many species, we will collect andanalyze some sample species to find the constant characters thatlead you to improve your knowledge about the groups and thefamilies.

Collecting
A good collection is an essential aspect of herbarium practice. A good herbarium
specimen must give the investigator the best possible information about the plant.
Theoretically and ideally, a specimen herbarium contains all parts of the plants (flowers
or fruits, leaves, wood, branches etc.) and should be well annotated so as to provide all
information not visible on the dry herbarium specimen. Many plants show leaf dimorphism;
plants are monoecious but the flower is unisexual (such as in Begoniaceae). To avoid
misinterpretation, it is recommended to always collect more than a single specimen.
As a general rule do not collect sterile material. For scientific purposes sterile herbarium
specimen are mostly useless, and for identification it is often very troublesome and timeconsuming.

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Small plants should be collected as a whole. Larger terrestrial herbs, shrubs, creepers
and small climbers can be easily collected with secateurs or pocket knife. When collecting
plant, please find a branch with leaves and flowers or fruits. Parts too large for mounting
on a herbarium sheet can be reduced in size before drying. The size of specimen should begood enough to fit between folded old newspapers. Collect a fair sized sample and not a
scrap.

Making Notes
Make it a habit not to rely on your memory but
to write your notes at the collecting site. Do not
wait till the end of the day, or worse still, to put it
off to the next day. After collecting your specimen,
give it a number tag and enter notes in yourcollection book under the corresponding number.
It is important to take note of all parts that may
disappear, or are invisible in the herbarium
specimen. Your field data of the living plantcollections will be transferred to the herbarium
label, which will then be placed on the corner ofthe sheet of herbarium collections, which will bear
the following information:a) Location (place of the collection, e.g., island,
province, district including altitude, latitude
and longitude) or a place easily located on amap.
b) Date (date of collection)c) Collecting number (person/s) who collected
the plant; the number has to be continuous; do not make new numbers for new
locations)d) Habitat (notes of vegetation type, topography, soil etc.)e) Habit (tree, shrub, liana, herbs)f) Exudate (latex, resin and its color, etc.)g) Color of various parts, such as flowers, fruit or young leafh) Ecological and biological conservation (phenology of flowers, pollination, seeddispersal)i) Local names (check if the local name is for a single plant or a group of plants, andconsistence)j) Local uses (as medicine, food, utensil etc., and how they are prepared)

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CYTOLOGY
Chromosome information is one of the important character information in taxonomy andplant systematics. Chromosome information is also very powerful in describing the mode of
reproduction (sexual, asexual, apomixis or hybrid), evolutionary biology, and relationshipamong species complex. Chromosome can be obtained during cell division at meiosis and/or mitosis of any single plant. Four main stages during cell division in plant are: Prophase,
Metaphase, Anaphase andTelophase. During cell division, at late metaphase stage, isusually the best time for chromosome counting, because during this stage the chromosome
is fully condensed into very short form. It is important to understand the process of cell
division during mitosis and meiosis.
Cell Divison in plant:
A. Prophase; B. Metaphase; C. Anaphase; D. Telophase

Chromosome information can be: 1)number of chromosome in eachcell; 2)size and morphology of any single chromosome; 3)reaction
to the staining; and 4)behavior of chromosome, especially duringmeiosis.

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Number of chromosomes
In somatic cell division during mitosis, a mother cell produces two daughter cells withthe same chromosome number. It is just a duplication of chromosome numbers. While ingametic cell division during meiosis, a mother cell produces four daughter cells with half
chromosome numbers. However, in apogamous plant, gametic chromosome number duringmeiosis produces the same chromosome numbers as mother cell. There is no cytokinesis at
the late meiosis.

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Size and morphology of chromosome
Size and morphology of chromosomes are important information during karyotype
analyses. Some time they are quite similar like in most of Pteridophyte, but some plants
have a great range in size and morphology like that of family Commelinaceae. Karyotype
analyses can demonstrate their relationship of chromosome morphology among species.

Size ofchromosome

Reaction to the staining and chromosome behavior
Chromosome behavior during meiosis is a good character for evaluating putative
hybrids. Hybrid plant exhibits irregular meiotic divisions, producing irregular-shaped andabortive spores or pollen.

Polyploidy
Not like in animals, polyploidy in plants is very common. Duplication of basic
chromosome numbers occurs often in any plant groups and ferns. The smallest
chromosome number in each plant species isusually treated as basic chromosome numberor (x). In gametic chromosome (n) it is half of the somatic chromosome (2n). The smallest
chromosome number (x) is usually treated as sexual diploid. Refer to chart below showing
the process of chromosome duplication in polyploid complex.

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Training Manual on Plants Taxonomy (Dicots) in Southeast Asia

Polyploid complex

Sources of chromosomes
Chromosome of plants can be observed in any active growing meristem, such as root
tips, young leaves, young bud of flowers, or young sporogenesis of ferns. Active vegetative
organs are produced by somatic cell division during mitosis,while active generative organs
are produced by gametic organs during meiosis.
Root tips: Select active root tip from seedling, or young plant in the gardens. Goodand healthy root tips are usually creamy in color.
Young leaves: Select good young leaves. They can be collected from plantcollections in the gardens, or any young leaves in the field.
Flower buds: Select young flower buds. For meiosis observation, select new flowers
in different stages of maturity. For mitosis observation, the treatment is like those ofyoung leaves.
Sporogenesis: For meiosis observation of fern group, select one young frond withdifferent stages of spore maturity.
Voucher specimens of any plant cytologically examined is very important.

PROCEDURE IN OBSERVING CHROMOSOMES
1. Observing somatic chromosomesin root tips
i. Pre-treatment (at 18-20O C)• Prepare a small bottle with 0.002 mole 8-hydroxyquinolin aqueous solution• Select good root tips, cut it at 1 cm long
• Put 4-6 good root tips into a solution bottle• Store this solution at 19O C (18O-20O C) for 3-5 hours
ii. Fixation at room temperature• Select root tips and clean it in clean water• Put it into a small bottle with 45% acetic acid
• Keep it at room temperature for 10 minutes

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iii. Maceration (at 60OC)• Prepare (1 NHCl : acetic acid = 3:1) in a small bottle• Put the bottle at 60O C boiling water• Keep it floating in boiling water for 2-4 minutes
iv. Staining at room temperature• Prepare 2 clean glass slides• Take 2 root tips from the solution and put it on glass slide, 1 root tip in eachglass slide• Cut 1-2 mm distal root tip and remove the other parts• Drop 1.5-2 % aceto-orcein into it• Store in a chamber containing wet filter paper with 45% acetic acid (5-10
minutes, no more than 1 hour)v. Observe using the microscope
2. Observing somatic chromosome in young leaves
i Pre-treatment (at 18-20O C)• Prepare a small bottle with (a): 0.02 – 0.05% aqueous solution of colchicines or(b): 0.002 mole 8-hydroxyquinolin aqueous solution or• Select good young leaves presumably with active mitosis• Store this solution at 19O C (18O-20O C)
ii. Fixation at ca 5OC• Put materials into modified Carnoy’s solution (absolute ethanol : glacial acetic
acid : chloroform = 2 : 1 : 1) for more than 1 hr, (storing it in a refrigator canlast a few months)iii. Maceration and staining• Put small part of young leaf on glass slide• Cut into smaller parts using sharpened needle• Drop 1.5-2% acetoorcein containing 1 N HCl (1/10 volume) and store in achamber containing wet filter paper with 45% acetic acid for several minutes.
iv. Tapping and squash• Move glass slide with the material from the chamber• Add one drop 45% acetic acid and cover it with a cover slip (root tip in thecenter of cover slip)• Warm up to 60OC for several seconds then put glass slide on a filter paper• Fix one corner of cover slip with a finger• Tap on cover slip gently using a sharpened stick or pencil by vertical direction
to set the sample and to remove bubbles• Put glass slide between folded filter paper, then squash gently but strongly• Seal cover slip with nail coat or paraffin
v. Observe using the microscope
3. Gametic chromosome observation in young flower
i. Fixation at 5O C
• 45 % acetic acid for more than 15 minutes• Modified Carnoy solution for more than 1 hour (move material into 45% acetic
acid just before use)ii. Staining• Put anthers on glass slide• Remove unnecessary part of anther from glass slide• Select pollen mother cells (PMCs) on glass slide (add fixative some time tokeep materials wet)
• Drop 1.5-2% acetoorcein and store in a chamber with filter paper wet withacetic acid for 2-3 minutes

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Training Manual on Plants Taxonomy (Dicots) in Southeast Asia

iii. Squash• Put glass slide between filter papers and squash it gently• Seal cover slip with nail coativ. Observe using the microscope
4. Gametic or meiotic chromosome observation in young frond of fern during
sporogenesis• Fix small portion of juvenile leaves with very young sporangia in glacial acetic
acid: absolute alcohol (1:3) solution.
• Keep at room temperature for 24 hours• Chromosome counting was made on SMCs at meiosis with aceto-carmine
squash method

HOW TO PREPARE SOLUTION
1). 0.002 mole 8-Hydroxyquinolin aqueous solution• 8-hydroxyquinolin (M.W.145.15)• 1 liter aquadest (145.15 x 0.002) = 0.29 g• 10 liter aquadest = 2.9 g• Shake and store for several days after all the powder has dissolved. Then,it canbe used for pretreatment(usually we need several days to dissolve the powder
completely).
2). 2% acetoorcein (total volume 50 cc)• Warm up 22.5 cc glacial acetic acid to 100O C using water bath
• Add 1 g orcein, then shake• Cool down to room temperature• Add 27.55 cc water, then filter• Place stock solution in colored bottle with filter
3).

Shiff’s solution (total volume 220 cc)• Boil 200 cc aquadesh (100Oc)filter and shake• Add 1 g fuchsin basic (caution on explosive boiling)• Or: add few grains of fuchsin basic and then add the remaining, or shake wellin boiled water and then add fuchsin basic• Cool down to ca. 500 C• Add 20 cc 1 N HCl• Cool down to room temperature• Add 1 g Sodium metabisulfate (Na2S2 O5) or potassium metabisulfite(K2S2O5) and shake for several minutes• Store at cool and dark place for 1 day• When solution becomes clear yellowish color, store inrefrigerator

MOLECULAR SYSTEMATICS LABORATORY WORKThe discipline of systematics study involves the diversity of organisms and theirevolutionary relationships among them. Systematics provides the essential framework
without which we could not recognize or study biological diversity and evolution.
Systematists use fossil, molecular, and genetic data to infer evolutionary relationships.
Molecular biology has revolutionized the field of Systematics. The DNA evolves by
mutations and these mutations are incorporated in the DNA in which they are fixed in
populations. This will lead to divergence of DNA sequences in different species. AlthoughTraining Manual on Plants Taxonomy (Dicots) in Southeast Asia

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diverged, we can refer to two DNA sequences as homologous (just as we would for any
morphological trait).
The steps in using molecular methods involve (1) DNA extraction; (2) gene amplificationor PCR (Polymerase Chain Reaction) and purification; (3) Cycle sequencing and purification;and (4) the DNA sequencing. Other than sequencing method, DNA fragments are also
used in molecular systematics. Upon obtaining the sequences or DNA fragment data,
phylogenetic analysis are carried out to interpret the relationships among the species
studied.
This manual is designed to introduce to beginners the theory and practice ofphylogenetic inference from molecular data (DNA sequences or fragments).

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What is DNA?
A chromosome consists of a DNA molecule packed together with proteins. The bacterial
chromosome is a double-stranded, circular DNA molecule associated with a small amountof protein,while Eukaryotic chromosomes have double-stranded linear DNA molecules
associated with a large amount of protein. Chromatinis a complex of DNA and protein andis found in the nucleus of eukaryotic cells. Histonesare proteins that are responsible for the
first level of DNA packing in chromatin.
Fig. 16-21b

Organization of DNA within chromosome(Source: Campbell 2008)

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Structures of DNA(Source: Campbell 2008)

Molecular Methods

Flowchart of Molecular Methods
DNA extraction
agarose gel

Gene amplification/PCR and purification
agarose gel

Cycle sequencing and purification
agarose gel

DNA sequencing

Phylogenetic analysis
Collecting Samples
There are some ways to collect samples for DNA extraction. The main principle toremember is to “kill” the cells as fast as possible to prevent any damage to the cells. If theliving plants are near your lab, you can put the samples right away in CTAB buffer placedin an Eppendorf tube and bring to the lab for further processing. Liquid nitrogen helps ingrinding the cells quickly and stopping any enzyme that will destroy the cells.
If you go to the field far away from the lab, the best method is to bring liquid nitrogen
if possible or put the samples (5 x 5 cm2 leaf cut in about 1 x 1 cm2) in a tea bag in silica

28

Training Manual on Plants Taxonomy (Dicots) in Southeast Asiagel inside a ziplock plastic bag. This would last some years, but secondary metabolites,
which prevent the PCR process, are often encountered. Extraction from fresh material isstill best so far for successful PCR process.
A kit is also available by scrubbing the samples on a card then it is punched andprocessed further according to the manual of instruction.

DNA Extraction from Plant Tissue
To extract DNA, several processes are involved, such as disrupting the cell; removal ofprotein, polysaccharides and other compounds; removal of CTAB; then finally precipitationof nucleic acid. There are several protocols that would suit best the samples we extract.
The simple one is explained here, modified from Doyle & Doyle 1987.
Flowchart of Processes in DNA Extraction (Doyle & Doyle 1987)

Cell disruption(mechanically by grinding, and chemically by 2x CTAB)

Removal of protein, polysaccharides, etc(with chloroform-isoamyl alcohol)

Removal of CTAB
(with wash buffer)

Precipitation of nucleic acids(with cold isopropanol)

DNA Extraction from peas, tomato and green split peas

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Protocol of DNA Extraction Modified from Doyle & Doyle 1987
1.
2.
3.
4.
5.
6.
7.

8.

9.
10.
11.
12.
13.
14.
15.
16.
17.
18.
19.
20.
21.
22.
23.

Label 1.5 ml Eppendorf tubes then place on a rack.
Prepare one circle cut out by punching fresh healthy leaves or silica gel driedmaterial using 1.5 ml Eppendorf tube lid per sample.
Add a pinch of sterile sand.
In another 1.5 ml Eppendorf tube (one tube is needed for each DNA sample),preheat 500 µl 2x CTAB with 0.2% mercaptoethanol (1 µl) at 65oC in a water bath.
Using forceps, submerge the tube that contains leaf sample in liquid nitrogen (ifany). If the lid is closed, openor pierce it.
Using a small plastic pestle, ground the tissue (sample) inside the tube to a fine dry
powder. Then, dissolve the powder in 500 µl 2xCTAB and add a pinch of PVP.
If there is no liquid nitrogen, add half of the preheated 2X CTAB (250 µl) to thetube containing the samples, then ground the tissue using a small plastic pestle to aslurry green state. Add 250 µl more of 2xCTAB and a pinch of PVP.
Place the tube containing the mixture in a vortex for a few seconds, then incubate
for an hour at 65oC in a water bath shaker to allow the cell to lyse for DNA
liberation.
Take out the tubes from the water bath shaker and coolfor 10 minutes.
Centrifuge the samples at 13,000 rpm for 10 minutes at 20oC.
Gently remove the aqueous (upper) phase without removing any of the particulatematter, then place in a clean 1.5 ml Eppendorf tube.
Add 500 µl chloroform: isoamyl alcohol (24:1) and mix well by inversion.
Place the samples on a shaker for 20 minutes.
Centrifuge the samples at 13,000 rpm for 10 minutes at 20oC.
Remove the supernatant gently, being careful not to pick up any of the bottom
layer, and place in a clean 1.5 ml eppendorf tube.
Repeat the above steps once again to re-extract the supernatant.
To precipitate the nucleic acids, add 2/3 volume (200-300 µl) freezer cold
isopropanol (propan-2-ol) then mix well by gentle inversion.
Continue the mixing by inversion until the oily appearance of the mixture is gone.
Leave the samples at-20oC overnight.
Centrifuge the samples at 13,000 rpm for 10 minutes at 20oC.
Pour off the supernatant, being careful not to pour out the pellet.
Dry the pellet in a vacuum drier for 10 minutes. Do not over-dry the pellet as it will
stick hard to the tube and will be difficult to dissolve.
Re-suspend the pellet to obtain the nucleic acid solution in 50 µl of TE buffer by
flicking the tube with a finger. The genomic nucleic acids can be stored in thefreezer until required.

Chemicals used and their function in DNA extraction process• 2x CTAB buffer contains CTAB, sodium chlorides, EDTA, beta-mercaptoethanol and
PVP.This extraction buffer is mostly to protect the DNA from degradation by native
enzymes and secondary plant metabolites.It is also a cationic detergent that helps tolyse the cell membranes and will form complexes with nucleic acids.
• Sodium chlorides help the formation of nucleic acid-CTAB complexes.
• EDTA chelates divalent ions, especially Ca2+ and Mg2+, and prevents the activity ofmetal-dependant nucleases.
• Beta-mercaptoethanol is a reducing agent that protects the DNA against quinones,
disulphites, peroxidases, and polyphenol oxidases.
• PVP will form complexes with secondary plant products, in particular, complex
polyphenols, tannins, and quinones.

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Training Manual on Plants Taxonomy (Dicots) in Southeast Asia

Visualizing DNA after extraction
The technique called electrophoresis is used to visualize DNA to check the quality andquantity of the DNA obtained either from extraction or PCR product. Each sample is run
on a 1% agarose gel (0.5 g agarosed) electrophoresis stained with Ethidium Bromide.

Agarose gel electrophoresis of DNA extraction productProtocol of gel electrophoresis
1. Heat the agarose in TBE or TAE buffer in a microwave until all particles are
dissolved. When boiling in the microwave, it is best to wait until the bubbles getbigger to avoid having small bubbles in the set gel.
2. When it is cooled, add 1 µl Ethidium Bromide then pour to a gel mould fitted with agel comb.
3. When the gel is set, remove the comb so that wells are formed.
4. Mix 5 µl of extracted total genomic DNA with 3 µl loading solution then load in awell.
5. Load 5 µl DNA size marker in one side well of the samples to compare with the total
genomic DNA.
6. Run in an electrophoretic field at 60-80V for 1-1.5 hours.
7. The negatively charged DNA will move to the positive electrode at a certain speed
which depends upon the size of the molecules. Observe the bands of ethidium
bromide corporated-DNA under UV light.
8. Document the results with a digital camera and print out.

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Agarose gel electrophoresis of PCR productHow to amplify DNA?
Polymerase Chain Reaction (PCR) to amplify DNA segment
Gene amplification to produce identical DNA copies is obtained via Polymerase Chain
Reaction (PCR) technique. Three basic stages are involved, i.e. denaturation, annealing
and synthesis or primer extension. The denaturation phase (high temperature) splits the
double-stranded DNA into single-stranded DNA. The annealing phase involves lowering
the temperature. In this phase, the oligonucleotide primers will bind to the single-stranded
DNA. The third stage, synthesis stage, involves the binding of polymerase enzyme (Taq=
Thermusaquaticuspolymerase) to deoxyribonucleotidetriphospates (dNTPs) and catalyze areaction by attaching the nucleotides to single-stranded DNA.

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Training Manual on Plants Taxonomy (Dicots) in Southeast Asia

PCR machine

Protocol of PCR
The PCR reaction mixtures of total volume of 50 µl in 0.2 ml PCR tube contains:• 34.5 µl sterile distilled water• µl of 2mM deoxyribonucleoside triphosphate (dNTP) mix• 5.0 µl of 10x reaction buffer (160 mM (NH4)2SO4, 670 mMTrisHCl pH 8.8 at 25oC,
0.1% Tween-20)• 2.5 µl of 50 mM MgCl2• 1.5 µl of 10 mM Primer 1• 1.5 µl of 10 mM Primer 2• 0.25 µl of 5U/µl thermostable DNA polymerase• 2 µl DNA template from aliquots of total genomic DNA
PCR cycle parameters for ITS (Internal Transcribed Spacer) amplification are as follows:• Initial denaturation for 3 minutes at 94oC
• Denaturation of template DNA for 1 minute at 94oC
• Primer annealing for 1 minute at 55oC
• Primer extension for 1.5 minute at 72oC
After 30 cycles, a final extension step of 5 minutes at 72oC is added. This extension ismeant to allow completion of unfinished strands.
Gel electrophoresis (method described previously) at 60-80 V for 1-1.5 hours using 1.5µl of PCR products will be carried out to check successful amplification and quantity of
PCR products. DNA size marker 100 bpladder is used for comparison of amplified DNA
obtained.

Purification of PCR ProductPurification means to purify the DNA obtained from any unwanted artefacts from PCR
such as primers, unincorporated nucleotide, polymerase and salts. There are several ways
to do purification:one is by adding polyethylene glykol or using ready kit, for example
QIAquick™ PCR Purification Kit. Follow the protocol provided by the kit if you use it.

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Cycle sequencing and purification of sequencing reactions
Protocol of cycle sequencing
Cycle sequencing is performed prior to automated sequencing. The 20 µl sequencing
PCR mixture contains:• 13 µl of sterile distilled water• 4 µl of Thermo Sequenase II reagent Premix• 1 µl of 5 mM of one primer type• 2 µl of DNA template (from purified PCR products)
The samples are then placed in a thermal cycler and run for 25 cycles with the following
PCR conditions:• Denaturation step for 10 seconds at 94oC
• Primer annealing for 5 seconds at 50oC
• Primer extension for 4 minutes at 60oC

Protocol of purification of sequencing reactions
The results of cycle sequencing are purified according to the following procedure.
• Transfer the 20 µl of PCR cycle sequencing products to a fresh 0.5 ml tube containing
2 µl of sodium acetate/EDTA buffer.
• Add 55 µl of 100% cold (-20oC) ethanol to each reaction.
• Mix briefly with a vortex mixer and place on ice for 15-20 min to precipitate theDNA.
• Centrifuge in a microcentrifuge for 15 minutes at 13,000 rpm.
• Remove the supernatant formed as much as possible.
• Add 250 µl of cold 70% ethanol to wash the pellet, then centrifuge at the same speed
for 5 min.
• Remove again as much as possible the supernatant formed.
• Vacuum-dry the pellet that remains at the bottom of the tube in a vacuum centrifuge
for 2-5 minutes.

Automated DNA Sequencing
Gel preparation and loading for automated DNA sequencing is performed on an ABI
automatic DNA Sequencer according to the manual supplied.

Electropherogram of DNA sequences

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Training Manual on Plants Taxonomy (Dicots) in Southeast Asia

PCR based techniques– nucleotide sequences– amino acids• synonymous (silent) vs. non-synonymous mutations– micro-satellites (few base pair repeats)– co-dominant markers• ‘DNA fingerprinting’• RFLP, AFLP, RAPD, ISSRSEQUENCE ANALYSIS
Sequence Editing
The sequence boundaries of each region studied are compared with the results ofexisting study or compared with the sequences in the gene bank. Each region should be
confirmed from forward and reverse sequences. There are several free programs to edit
sequences such as 4Peaks for Macintosh or BioEdit for Windows. Licensed programs withadditional function to assemble the complementary strands automatically from forward andreverse sequences are available such as Chromas, Genious, etc. These programs can build aconsensus sequences. Ambigious bases are edited manually after observing the bases fromforward and reverse sequences.

Sequence alignment
Sequence alignment is carried out using Clustal program. Other program called Muscle
is also powerful for sequence alignment. These alignments are subsequently refined by eye.

Alignment of DNA Sequences

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PHYLOGENETIC ANALYSIS
Phylogeny and the Tree of life
Phylogeny is the evolutionary history of a species or group of related species.
WilliHennig (1913-1976) introducedphylogenetic systematic. In 1950, he wrote
GrundzügeeinerTheorie derPhylogenetischenSystematik. It was in 1966 that Phylogenetic
Systematics became known.
Phylogeny is based upon evolutionary relationships, i.e., upon common ancestry. A
cladogram is a tree diagram which depicts hypothesised evolutionary history, while aphylogram is a tree which indicates by branch length the degree of change believed to haveoccurred along each lineage.

A phylogenetic tree represents a hypothesis about evolutionary relationships. Eachbranchpointrepresents the divergence of two species. Sister taxaare groups that share an
immediate common ancestor.
A rootedtree includes a branch to represent the last common ancestor of all taxa in thetree. A polytomyis a branch from which more than two groups emerge. Phylogenies areinferred from morphological and molecular data. To infer phylogenies, systematists gather
information about morphologies, genes, and biochemistry of living organisms.

Morphological and Molecular Homologies
Organisms with similar morphologies or DNA sequences are likely to be more closely
related than organisms with different structures or sequences. When constructing aphylogeny, systematists need to distinguish whether a similarity is the result of homology
or analogy. Homology is similarity due to shared ancestry. Analogy is similarity due toconvergent evolution.

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Training Manual on Plants Taxonomy (Dicots) in Southeast Asia

Cladistics-Example

Evaluating Molecular Homologies
Systematists use computer programs and mathematical tools when analyzing comparable
DNA segments from different organisms.

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37

It is also important to distinguish homology from analogy in molecular similarities.
Mathematical tools help to identify molecular homoplasies, or coincidences. Molecularsystematics uses DNA and other molecular data to determine evolutionary relationships.
Once homologous characters have been identified, they can be used to infer a phylogeny.

Cladistics
Cladistics group organisms by common descent.A clade is a group of species that
includes an ancestral species and all its descendants. Clades can be nested in larger clades,
but not all groupings of organisms qualify as clades. A valid clade is monophyletic,
signifying that it consists of the ancestor species and all its descendants.

A valid clade is monophyletic, signifying that it consists of the ancestor species and allits descendants.
A paraphyletic grouping consists of an ancestral species and some, but not all, of thedescendants.
A polyphyletic grouping consists of various species that lack a common ancestor.

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Training Manual on Plants Taxonomy (Dicots) in Southeast Asia

39

Computer Software Packages Available for Data Analysis
Phylogeny Programs – Joe Felsenstein’s site lists 387 phylogeny reconstruction packages
and 28 free servers (Department of Genome Sciences and the Department of Biology at the
University of Washington)
PAUP* 4.0 – Phylogenetic Analysis Using Parsimony and other Methods
PHYLIP – package of programs for inferring phylogenies
MacClade – A useful software package for Phylogenetic Analysis
Mesquite – a modular system for evolutionary analysis, by W. P. Maddison and D. R.
MaddisonHennig86 – A PC-DOS program for phylogenetic analysis
MEGA – Molecular Evolutionary Genetics Analysis. MEGA is a DOS program for analyzing
molecular data. Developed by Sudhir Kumar, Koichiro Tamura and Masatoshi Nei (1993).
MrBayes – is a program for constructing phylogenetic trees by Bayesian method(Huelsenbeck 2000)
DAMBE – Data Analysis in Molecular Biology and Evolution, developed by X. Xia at the
University of Hong Kong
LVB – Reconstructing evolution with parsimony and simulated annealing
Other Programs – Maintained by Dr. Joe Felsenstein, Department of Genetics, The
University of Washington
Clann– Constructing Supertrees from partially-overlapping datasets, designed and written
by Chris Creevey at the Bioinformatics and Pharmacogenomics Laboratory at NUI MaynoothSuperTree software by Nicolas Salamin, Biophore - 1015 Lausanne - Switzerland
SuperTree Server by Mike Sanderson, D. Gusfield, and Oliver Eulenstein, Computational
Biology Laboratory, Department of Computer Science Iowa State University, Ames, IA
SuperTree software by Roderic D. M. Page, DEEB, IBLS, University of Glasgow, Glasgow
G12 8QQ, United Kingdom
TreeBASE by Bill Piel, M. Donoghue and Mike Sanderson. TreeBASE is a relational
database of phylogenetic information hosted by the Yale Peabody Museum

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Training Manual on Plants Taxonomy (Dicots) in Southeast Asia

APPENDIX

1. Directory of Participants to the Training Workshop on Taxonomy of Terrestrial Plants
2. Training Course Programme
3. Lectures and Powerpoint presentations during the training course
4. Group Reports
5. Pre- and Post-Training Evaluation forms
APPENDIX 1 – Directory of Participants to the Training Workshop on Taxonomy of
Terrestrial Plants
PARTICIPANTS
Name

Institution

Y. Lavy
ylavy21@yahoo.com

Deputy Director of Dept. of Wetland andCoastal Zone
Ministry of Environment
Cambodia

KethSamath
samathketh@yahoo.com

Researcher and Seed Production of Vegetable
General Directorate of Agriculture
Ministry of Agriculture, Forestry and Fisheries
Cambodia

TulusPambudi

BetungKerihun National Park
Kalimantan Barat
Indonesia

Tri Arfiantitri.arfianti@gmail.com

Purwodadi Botanical Garden - LIPI
East Java
Indonesia

Nor Ezzawanis Binti Abdullah Thaniezzawanis@frim.gov.my

Research Officer
Forest Research Division
Forest Research Institute of Malaysia
52109 Kepong, Selangor
Malaysia

Jovita Elderson Anak Ripen
jovita@sbc.org

Research Officer
Serawak Biodiversity Centre
KM20 Jalan Borneo Heights, Semengoh,
Locked Bag no. 3032, 93990
Kuching, Sarawak
Malaysia

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42

Name

Institution

KhinMyoThet
kmthetuof@gmail.com

Demonstrator
University of Forestry, Forest Department
Ministry of Forestry University of Forestry
Yezin, Nay Pyi Taw
Myanmar

Cecille G. Garcia
cgarcia519@yahoo.com

Ecosystem Management Specialist I
Protected Areas and Wildlife Bureau
Department of Environment andNatural Resources
Philippines

DhiocelA. Celadina
dhiocel@gmail.com

Instructor I
2nd Floor Faculty Room CFESWPU
Western Philippines University
San Juan Aborlan, Palawan
Philippines

Yow Wei Chingweichingyow@gmail.com

Research AssistantCenter for Tropical Forest Science 329
Jurong East Ave 1, #09-1690, Jurong 600329
Singapore

Tan SiuYueh
jon86nomad@yahoo.com.sg

Research Assistant
Plant Systematic Lab.
Department of Biological Sciences
National University
Singapore

BordintornSonsupabt.um.bdt@hotmail.com

Agricultural Scientist
Bangkok Herbarium
Department of Agriculture
50 Phaholyothin Rd. Chatuchak, Bangkok 10900
ThailandJarearnsak Sae Wei
wei_zhiling@yahoo.com

Researcher
Prince of Songkhla University
Herbarium, Princess Maha Chakri Sirindhorn
Natural History Museum Department of Biology,
Faculty of Science
Prince of Songkhla University, Hat Yai,Songkhla, 90112
ThailandNguyen The Cuong
cuongntc1979@yahoo.com

Researcher
Institute of Ecology and Biological Resources
Vietnam Academy of Science and Technology
18 Hoang Quoc Viet-CauGiay- Ha Noi
VietnamTraining Manual on Plants Taxonomy (Dicots) in Southeast Asia

NameInstitution

Nghiem Thi Phuong Le
nghphuongle@gmail.com

Biodiversity Conservation Agency
Vietnam Environment Administration
Ministry of Natural Resources and Environment
5th Floor, 99 Le Duan Str., Ha Noi
Viet Nam

Taufikurrahman Nasution

Cibodas Botanical Garden - LIPIWest Java
Indonesia

Indah Wahyuni
iendah_mail@yahoo.com

Research Assistant
SEAMEO BIOTROP
Jl. Raya Tajur Km.6 PO Box 116, Bogor
Indonesia

Lulut Dwi Sulistyaningsih
herbogor@indo.net.id

Herbarium Bogoriense, Botany Division
Research Center for Biology -LIPI, Cibinong
Indonesia

Bing Liu
liu_bing5858@163.com

Institute of Botany
Chinese Academy of Sciences
China

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RESOURCE SPEAKERS

44

Country

Name

Designation/Organization/ Contact Information

Japan

Shinji Fujii

University of Human Environments
Motojyuku, Okazaki 444-3505
Japan
Fax: +81 564 48 7814
shinji@uhe.ac.jp

Japan

Eiji Suzuki

Professor, Earth and Environmental Sciences
Graduate School of Science and Engineering,Kagoshima University
Tel.: +81 99 285 8942
Fax: +81 99 285 8942
suzuki@sci.kagoshima-u.ac.jp

Philippines

Edwino Fernando

Professor, University of the Philippines –Los Baños
Department of Forest Biological Sciences, UPLB,
College, Los Banos, Laguna 4031 Philippines
Tel.: +63 49 5362773
Fax: +63 49 5362773
Mobile: +63 917 5017153
edwino_fernando@yahoo.com.phIndonesia

DedyDarnaedi

Senior Researcher
Herbarium Bogoriense, Research Center for
Biology-LIPI
JlKomkpleks LIPI, Baranang Siang B-118 Town/
City: Bogor 16122 West Java, Indonesia
Tel: +62 251 8322859
Mobile: +62 08 1111 3420
darnaedi@yahoo.com or dedy@indo.net.id

Indonesia

TeguhTriono

tt_rekso@yahoo.com

Indonesia

Rugayahtitikrugayah@yahoo.com

Indonesia

Harry Wiriandinata

harry_wiria@yahoo.com

Indonesia

MarlinaArdiyani

marlina.ardiyani@googlemail.com

Indonesia

Nanda Utami

utami_16002@yahoo.com

Training Manual on Plants Taxonomy (Dicots) in Southeast Asia

APPENDIX 2 – Training Course Programme

TRAINING-WORKSHOP ON PLANT TAXONOMY
RC Biology LIPI - Indonesia, 16 - 23 February 201115/02/2011
Day 1

Day 2

16/02/2011

17/02/2011

Participants arrival at
CSC, Indonesia
08.00 -09.00

Registration ofparticipantsCommittee

LMR

09.00-09.30

Opening Ceremony
Welcome remarks
Opening Message

MC: Wita Wardani
LIPI: Prof. Dr. Bambang
Prasetya
ACB/JAIF: Dr. Clarissa Arida
JWRC/MoE Japan:
Dr. Masaaki Yoneda
MoE Indonesia: Dr. Arief
JuwonoLMR

09.30-10.00

Introduction ofparticipants
Overview of the training
Workshop(Photo opportunity)

Dr. Dedy Darnaedi
JAIF-ACB Project
ESABII-GTI-JWRC

LMR

10.00-10.30

Break

10.30-12.00

General Lecture on Plant
TaxonomyTT

LMR

12.00-13.00

Lunch

13.00-15.00

General Lecture on
Biogeography
and lecture on Ericaceae

15.00-15.20

Break

15.20-17.00

Visit Herbarium
Collection

08.00-08.30

Registration ofparticipants

08.30-10.00

General lecture on Plant
Morphology of
Dicotyledoneae
and lecture on
Sapotaceae

10.00-10.30

Break

10.30-12.00

Lecture on Plant Age

12.00-13.00

Lunch

13.00-15.00

Lecture on Begoniaceae,
Fabaceae,
Rosaceae&Rubiaceae

15.00-15.30

Break

15.30-16.15

Lecture on
Cucurbitaceae

R

LMR

16.15-17.00

Lecture on
Balsaminaceae

NU

LMR

SMR
EF

LMR
LMR

HW

LMR
LMR

TT

LMR

LMR
ES

LMR
SMR

HW

LMR

LMR

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45

Day 3

Day 4

Day 5

Day 6

46

18/02/2011

19/02/2011

20/02/2011

21/02/2011

08.00-08.30

Registration ofparticipants

08.30-10.00

Lecture on Cytology

10.00-10.30

Break

10.30-12.00

Lecture on Molecular
Systematic

12.00-13.00

Lunch

13.00-15.00

Lecture on Database IBIS
and visit Database Room

15.00-15.30

Break

15.30-17.00

Working with herbarium
specimens

08.00-08.30

Registration ofparticipants

08.30-12.00

Working on Cytology(group I)
Working on Molecular
Analyses (group II)

12.00-13.00

Lunch

13.00-16.30

Working on Cytology(group II)
Working on Molecular
Analyses (group I)

08.00-08.30

Registration ofparticipants

08.30-10.00

Lecture on Ecology andForest Dynamic

10.00-10.30

Break

10.30-12.00

Sampling method

12.00-13.00

Lunch

13.00-15.00

Lecture on Fagaceae

15.00-15.30

Break

15.30-17.00

Preparation for field trip

06.30-07.00

Registration ofparticipants

07.00-09.00

Move to Cibodas Bot.
Gard.

09.00-10.00

Break

10.00-15.00

Visit Gunung GedePangrango National
Park
and Cibodas Botanical
Garden

LMR
DD

LMR
LMR

MA

LMR
SMR

BH/BBM

LMR &DBR

HW, R, NU

BSLDD, UHLC

MA, WW, YS

LMS
SMR

DD, UHLC

MA, WW, YS

LMS
LMR

ES

LMR
LMR

ES, HW

LMR
SMR

SF

LMR
LMR

ES, HW

LMR
LMR

Committee

CBG
CBG

Committee

GGNP
CBG

15.00-15.30

Break

15.30-17.00

Data Analysis

ES

CBG

17.00-21.00

Dinner & Discussion

Committee

CBG

Training Manual on Plants Taxonomy (Dicots) in Southeast Asia

CBG

Day 7Day 8

22/02/2011

23/02/2011

07.00-08.00

Breakfast

CBG

08.00 -12.00

Sample collections &Processing herbarium

12.00-13.00

Lunch and packing

13.00-16.00

Back to CSC

16.00 -......

Preparing report andpresentation (eachgroup)

08.00-08.30

Registration ofparticipants

LMR

08.30-10.00

Preparing report andpresentation (eachgroup)

LMR

10.00.10.30

Break

LMR

10.30-12.00

Preparing report andpresentation (eachgroup)

LMR

12.00-13.00

Lunch

SMR

13.00-15.00

Group presentation anddiscussion

LMR

15.00-15.30

Break

LMR

15.30-17.00

Feed back fromparticipants

LMR

Closing remarks:

LMR

CBG
CBG

Committee

JWRC
ACB
LIPI :
Director of RC Biology
24/02/2011

HW

CBG

Departure

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