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P di f Proceedings of gBiotechnology International ConferenceBiotechnology International Conference
i h l f if“Biotechnology for Better Life”gy
InstitutTeknologi Bandung15‐16 June 2009
PREFACEPREFACE
This CD contains the abstracts of the talks presented in Biotechnology International Conference (BIC) with an This CD contains the abstracts of the talks presented in Biotechnology International Conference (BIC) with an overall theme on “Energy and the Environment: Reinvention for Developing Countries”, especially overall theme on Energy and the Environment: Reinvention for Developing Countries , especially “Biotechnology: Biotechnology for Better Life”. This is in line with commemorating the 50th anniversary of ITB Biotechnology: Biotechnology for Better Life . This is in line with commemorating the 50 anniversary of ITB (InstitutTeknologi Bandung) as one of the oldest education and science institution in Indonesia. (InstitutTeknologi Bandung) as one of the oldest education and science institution in Indonesia.
Bi h l I i l C f h ld i W H ll (A l B ) d E H ll (A l Ti ) Biotechnology International Conference 2009 was held in West Hall (Aula Barat) and East Hall (Aula Timur) at I tit t T k l i B d th 6th f J Th BIC i t t th d l t f InstitutTeknologi Bandung on 15th‐16th of June 2009. The BIC aims to promote the development of bi t h l d th i l i th d l t f i d t h l i I d i d R i l biotechnology and their roles in the development of science and technology in Indonesia and Regional.
There are three areas of research covered in this conference. They are biotechnology of human health, y gy ,biotechnology of industrial and applications, and biotechnology of agricultural. The topics of research were gy pp , gy g ppresented in six invited speakers and 34 contributed talks. We would like to thank the invited speakers and p p pthose who have submitted papers to this conference.
We wish you a enjoy conference and stay in BandungWe wish you a enjoy conference and stay in Bandung.
Bandung, 10th of June 2009g, 9On behalf of the Organizing Committee,g g ,
Adi PancoroAdi PancoroDirector of Biotechnology Research Center ‐ ITBDirector of Biotechnology Research Center ITB
SCIENTIFIC COMMITTEESCIENTIFIC COMMITTEE
Prof. Dr. Tjandra Setiadi, Bandung Institute of Technology, Indonesia
Prof. Dr. Wisjnuprapto, Bandung Institute of Technology, Indonesia
Dr.Achmad Saifudin Noer, Bandung Institute of Technology, Indonesiag f gy
Dr. Adi Pancoro, Bandung Institute of Technology, Indonesia, g f gy,
Dr. Akhmaloka, Bandung Institute of Technology, Indonesia, g f gy,
Dr. Anggraini Barlian, Bandung Institute of Technology, IndonesiaDr. Anggraini Barlian, Bandung Institute of Technology, Indonesia
Dr Edwan Kardena Bandung Institute of Technology IndonesiaDr. Edwan Kardena, Bandung Institute of Technology, Indonesia
Dr Enny Ratnaningsih Bandung Institute of Technology IndonesiaDr. Enny Ratnaningsih, Bandung Institute of Technology, Indonesia
Dr Ernawati A Bandung Institute of Technology IndonesiaDr. Ernawati A., Bandung Institute of Technology, Indonesia
Dr Fida MadayantiW Bandung Institute of Technology IndonesiaDr. Fida MadayantiW., Bandung Institute of Technology, Indonesia
D I G W t B d I tit t f T h l I d iDr. I.G. Wenten, Bandung Institute of Technology, Indonesia
D M li R d i M i B d I i f T h l I d iDr. Maelita Ramdani Moies, Bandung Institute of Technology, Indonesia
l d f h l dDr. Marselina IrasoniaTan, Bandung Institute of Technology, Indonesia
Dr. Debbie S. Retnoningrum, Bandung Institute of Technology, Indonesia
Dr. Dessy Natalia, Bandung Institute of Technology, Indonesia
Dr. Elfahmi, Bandung Institute of Technology, Indonesia
Dr. Ronny Purwadi, Bandung Institute of Technology, Indonesia
Dr. Sony H. Sumarsono, Bandung Institute of Technology, Indonesiay , g f gy,
Dr. Sony Suhandono, Bandung Institute of Technology, Indonesiay , g f gy,
Dr. Sri Nanan B. Widiyanto, Bandung Institute of Technology, IndonesiaS a a d ya to, a du g st tute of ec ology, do es a
Dr. Zeily Nurachman, Bandung Institute of Technology, IndonesiaDr. Zeily Nurachman, Bandung Institute of Technology, Indonesia
CONTENTSCONTENTS
A. BIOTECHNOLOGY OF HUMAN HEALTHA. BIOTECHNOLOGY OF HUMAN HEALTH
Antioxidant and Antibacterial Activities of Extracts from Hoan Ngoc (Pseuderatherum Platiferum) a Medicinal PlantAntioxidant and Antibacterial Activities of Extracts from Hoan Ngoc (Pseuderatherum Platiferum), a Medicinal PlantChantana KankamolChantana Kankamol
Humanization of MurineAnti CEA antibody and the Selection of High Antibody Producing Cell Lines Using a Rapid Humanization of MurineAnti‐CEA antibody and the Selection of High Antibody‐Producing Cell Lines Using a Rapid Automated SystemAutomated SystemHeilly Chong, Ong Sheau Ching, Suba Dharshanan, Siong Jong Hang, Khairul Ezani Najamudin, Zulkeflie Zamrod
Single Amino Acid Residue Substitution in Two Isolates of Isoniazid‐Resistant Mycobacterium tuberculosis Catalase‐g yPeroxidasePurkan, Ihsanawati, Debby Soffie Retnoningrum, Dessy Natalia, and Achmad Saifuddin Noer
Analysis of Alpha‐Glucosidase Inhibitors Extracted From Several Medicinal HerbsStephanie, Elisa Friska Romasi, Maggy Thenawidjaja
From Science to Farming and the Seek for the Future Policy on Agricultural Biotechnology Product Commercializationl dRagil Yoga Edi
DNA Methylation on Rar Β2 Promotor Is Epigenetic Process on Breast CancerI K t t G dh M li th Wibi Ri A h Hid ti d Fit i A iI Ketut Gedhe Muliartha, Wibi Riawan, Asnah Hidayati and Fitri Armania
Identification of a Calcium‐Dependent Protein KinaseGene Involved in Osmoregulation in Chlamydomonas reinhardtiiReinhard PinontoanReinhard Pinontoan
Mi h d i l DNA Di i i I di P l i f S h P d i i li i N i P Mitochondrial DNA Diversity in Indigenous Populations of Southern Papua and its implications on Native Papuan HaplogroupsHaplogroupsRichardo Ubyaan, Epiphani I. Y. Palit, Hendry Yanuar, Yohanis NgiliRichardo Ubyaan, Epiphani I. Y. Palit, Hendry Yanuar, Yohanis Ngili
The Role Expression and Production of Brain Receptor Protein of Humpback grouper Cromileptes altivelis to Viral The Role, Expression and Production of Brain Receptor Protein of Humpback grouper Cromileptes altivelis to Viral Nervous Necrotic Infection for Development of Diagnostic MaterialNervous Necrotic Infection for Development of Diagnostic MaterialUun Yanuhar
Nucleotide Polymorphism on D‐Loop Human Mitochondrial DNA from Endoderm, Mesoderm, and Ectodermy p p , ,Yohanis Ngili, A. Saifuddin Noer, Adang Suwandi Ahmad, and Yoni F. Syukriani
B. BIOTECHNOLOGY FOR INDUSTRIAL AND APPLICATIONB. BIOTECHNOLOGY FOR INDUSTRIAL AND APPLICATION
Earth to Space: The Adventure of T1 LipaseEarth to Space: The Adventure of T1 LipaseAbu Bakar Salleh Thean Chor Leow Raja Noor Zaliha Raja Abdul Rahman and Mahiran BasriAbu Bakar Salleh, Thean Chor Leow, Raja Noor Zaliha Raja Abdul Rahman, and Mahiran Basri
Design of an Enzymatic Membrane Microreactor and Its Aplication for Transesterification ReactionDesign of an Enzymatic Membrane Microreactor and Its Aplication for Transesterification ReactionAchmadin Luthfi, Siswa Setyahadi, Misri Gozan, Mohammad Nasikinc ad ut f , S s a Setya ad , s Go a , o a ad as k
Xylanase Production from Winter Mushroom Flamulina velutipes Grown on Optimized Cultural ConditionsXylanase Production from Winter Mushroom Flamulina velutipes Grown on Optimized Cultural ConditionsAltaf Simairf
The Prospective of Raw Material and Pathway Modification in Polylactic Acid Production as Ecofriendly The Prospective of Raw Material and Pathway Modification in Polylactic Acid Production as Ecofriendly Biodegradable PlasticgDiana Lo
Ethanol Production from Glycerol of Biodiesel Plant using Batch and Semi‐continuous Fermentation by Enterobacter gaerogenes TISTR 1468kb k k k hAkbar Ciptanto, Poonsuk Prasertsan, Aran Hanpongkittikun, Sompong O‐Thong
Biodegradable Plastic Made from Hidrocoloid Alginate Extracted from Sargasum filifendulaM di hMurdinah
Solid State Fermentation for Improvement of Fatty Acids in By ProductsMohammad Faseleh JahromiMohammad Faseleh Jahromi
C i f W f Bi di l I d i V l bl P d I i i l S Bi fiConversion of Waste from Biodiesel Industry into Valuable Products: an Initial Step to BiorefineryAndi Ronny Purwadi M T A P Kresnowati and Tjandra SetiadiAndi, Ronny Purwadi, M.T.A.P. Kresnowati, and Tjandra Setiadi
……
Extraction and Characterization of Protease from The Viscera of Skipjack Tuna Fish (Katsuwonus pelamis Linnaeus)Extraction and Characterization of Protease from The Viscera of Skipjack Tuna Fish (Katsuwonus pelamis Linnaeus)Mery Tambaria Damanik Ambarita Jeni Anggrek Maggy ThenawidjajaMery Tambaria Damanik Ambarita, Jeni Anggrek, Maggy Thenawidjaja
Mechanism of Interaction between Prefoldin and Group II ChaperoninMechanism of Interaction between Prefoldin and Group II ChaperoninMuhamad Sahlan Taro Kanzaki Phan The Tai Tomatsu Zako Mizuo Maeda and Masafumi YohdaMuhamad Sahlan, Taro Kanzaki, Phan The Tai, Tomatsu Zako, Mizuo Maeda and Masafumi Yohda
Cloning of a Soft Coral Associated Bacillus aquimaris Α Amylase GeneCloning of a Soft Coral Associated Bacillus aquimaris Α‐Amylase GeneF Puspasari, D Natalia, Z Nurachman, O K Radjasa, A S NoerF. Puspasari, D. Natalia, Z. Nurachman, O. K. Radjasa, A. S. Noer
Effects of Temperature and pH on the Capability of Enterobacter aerogenes as Hg remediatorsEffects of Temperature and pH on the Capability of Enterobacter aerogenes as Hg remediatorsLanny Hartanti, Oktavina Maharani and Dien A. Limyatiy , y
Lignin recovery from pulp and paper mill wastewaterLignin recovery from pulp and paper mill wastewaterSandhya Babel, Massalin Suksai and Phattaraporn Sarikkha
Isolation, purification and characterization of cellulase from giant snail (Achatina fulica) and application to hydrolyze , p g ( ) pp y ycellulolytic materialsSendy Adhika Noor and Aran H‐Kittikun
The prospect of gadung (Dioscorea hispida Dennst.) tuber as carbon source for Aspergillus niger in amylase productionS Ri R t ih W d Nil bSuraya, Rina Ratnasih, Wardono. Niloperbowo
i id l i i filNitrous Oxide Gas Removal Using Compost BiofilterTania Surya Utami Josia Simanjuntak Heri Hermansyah Slamet Mohammad NasikinTania Surya Utami, Josia Simanjuntak, Heri Hermansyah, Slamet, Mohammad Nasikin
C. BIOTECHNOLOGY FOR AGRICULTURAL PRODUCTIONC. BIOTECHNOLOGY FOR AGRICULTURAL PRODUCTION
Expression of Nucleoprotein Gene of Newcastle Disease Virus and Its Utility in Seroprofiling of the Disease in Expression of Nucleoprotein Gene of Newcastle Disease Virus and Its Utility in Seroprofiling of the Disease in ChickenChickenVinod Kumar
Production and Application of Inoculant Acid‐Alumunium Tolerant Bradyrhizobium japonicum Strains with SoybeanProduction and Application of Inoculant Acid Alumunium Tolerant Bradyrhizobium japonicum Strains with SoybeanNisa Rachmania Mubarik
Detection and Cloning of A Gene Involved in Zwitermicin A Biosynthesis from Plant Growth Promoting g y gRhizobacterium Bacillus sp CR64Rika Indri Astuti, Aris Tri Wahyudi, Sarah Asih Faulina, and Nisa Rachmania
Analysis Polymorphism Gene in Cattle by DNA SequencingTety Hartatik, Tri Satya Mastuti Widi and Mohammad Cahyadi
Neuroethics: The Pros and ConshPriya Chatteree
Proceedings ofgBiotechnology International Conference“Biotechnology for Better Life”ISBN : 978-979-19994-0-3
Additional copies of this publication are available from:ITB Research Center on Biotechnology (PP Bioteknologi ITB)nd fl I t U i it C t (PAU B ildi )2nd floor, Inter University Center (PAU Building),
Institut Teknologi BandungInstitut Teknologi BandungJl. Ganesha no. 10, Bandung, IndonesiaPhone : +62‐22‐253 4115F 6Fax : +62‐22‐253 4115
The ITB’s Golden Anniversary
International Conference on
Energy and the Environment: Reinvention for Developing Countries
Bandung, 15th-20
th of June 2009
Biotechnology for a Better Life
Day 1: June 15 / Monday
Venue: West Hall
08:00-08:30 Registration of Participants
08:30-09:30 Opening Ceremony Rector of ITB Keynote: Economic Challenging in the Next Five Year
Tentative Keynote:
09.00-09.30 Keynote Speech National Research Council and its Roles in Solving Problems and Needs in Developing
World by Prof. Dr. Ir. Andrianto Handojo (DRN) (*) ** Ibu Tusi Adibroto
09.30-10.00 Invited Speaker Earthquake by Prof. Sri Widiyantoro (ITB)
10.00-10.30 C O F F E E B R E A K
Session PLENARY (Chairman : Dr. Achmad Saifudin Noer)
10.30-11.00 Invited Speaker Current and Future Trends of Therapeutic Proteins by Dr. Debbie S Retnoningrum (School of Pharmacy-ITB)
11.00-11.30 Invited Speaker Current Status of Biotechnology in Agriculture by Tumpal Silalahi (PT Monsanto-Indonesia).
11.30-12.00 Invited Speaker Value Added Products from Palm Oil mill by enzyme technology by Prof. Aran H-Kittikun (Prince of Songkla University Thailand).
12.00-13.00 L U N C H B R E A K
13.00-15.00 Technical Session 1 by Chairman: - (in detailed agenda)
15.00-15.30 C O F F E E B R E A K
15.30-17.00 Technical Session 5 by Chairman: - (in detailed agenda)
17.00-19.00 PREPARATION TO DINNER
Venue: East Hall 13.00-15.00 Technical Session 2 by Chairman: -
(in detailed agenda)
Technical Session 3 by Chairman: -
(in detailed agenda)
Technical Session 4 by Chairman:
- (in detailed agenda)
15.00-15.30 C O F F E E B R E A K
15.30-17.00 Technical Session 6 by Chairman: - (in detailed agenda)
Technical Session 7 by Chairman: - (in detailed agenda)
Technical Session 8 by Chairman: -(in detailed agenda)
17.00-19.00 PREPARATION TO DINNER
Day 2: June 16 / Tuesday
Venue: East Hall
08.30-10.00 Technical Session 9 by Chairman:- (in detailed agenda)
Technical Session 10 by Chairman: - (in detailed agenda)
Technical Session 11 by Chairman: - (in detailed agenda)
10.00-10.30 C O F F E E B R E A K
Session PLENARY (Chairman : Dr.Ronny Purwadi )
10.30-11.30 Invited Speaker Toward Self Sufficient in Food and Biopharmaceutical Through Biotechnology by Dr. Sony Suhandono (School of Life Sciences and Technology – ITB).
11.00-11.30 Invited Speaker Marine Biotechnology in Indonesia : Present status and future Development by Prof. Dr. Hari Eko Irianto (Director, Research Center for Marine and Fisheries
product Processing and Biotechnology, Ministry of Marine Affairs and Fisheries).
11.30-12.00 Invited Speaker Overview of Current Status and Development Strategy of Biotechnology in Indonesia by Dr. Bambang Prasetya (Head of Biotechnology Consortium Indonesia).
12.00-13.00 L U N C H B R E A K
Venue: West Hall 13.00-15.00 MRA FORUM for Biotechnology. Chairman: Dr. Adi Pancoro
15.00-15.30 C O F F E E B R E A K
15.30-16.30 MRA FORUM for Biotechnology. Chairman: Dr. Adi Pancoro
16.30-17.00 Closing
17.00-19.00 PREPARATION TO DINNER
The ITB’s Golden Anniversary
International Conference on
Energy and the Environment: Reinvention for Developing Countries
Bandung, 15th-20
th of June 2009
Day 1 : MONDAY, JUNE 15th 2009
TIME PARALLEL SECTION
VENUE : WEST HALL VENUE : EAST HALL - 1 VENUE : EAST HALL - 2 VENUE : EAST HALL - 3
Chairman: Dr. Sony H.
Sumarsono
Chairman: Dr. Zeily
Nurachman
Chairman: Dr. Fida
Madayanti W
Chairwoman: Dr. Sri Nanan
B. Widiyanto
Biotechnology of Human
Health
Biotechnology of Industrial
and Application
Biotechnology of Industrial
and Application
Biotechnology of Agricultural
Productions
13.00 :13.30 A.1 Chantana Kankamol
Antioxidant and antibacterial
activities of extracts from
Hoan Ngoc (Pseuderatherum
platiferum), a medicinal plant
B.1 Abu Bakar Salleh
Earth To Space: the
Adventure of Ti Lipase
B.8 Tjandra Setiadi
Conversion of waste from
Biodiesel Industry into
Valuable: an Initial step to
Biorefinery
C. 1 Vinod Kumar Sahu
Expression of nucleoprotein
gene of Newcastle disease virus
and its utility in seroprofiling of
the disease in chicken
13.30 :14.00 A.2 Zulkeflie Zamrod
Humanization of Murine
Anti-CEA and Selection of
High Antibody Producing
Cell Lines Using A Rapid
Automated Clonal Selection
Method
B.2 Achmadin Luthfi
Design Of An Enzymatic
Membrane Microreactor And
ITS Application for
Transesterification Reaction
B.9 Mery Tambaria
Damanik Ambarita
Extraction and
Characterization of Potease
from the Viscera of Skipjack
Tuna Fish(Katsuwonus
pelamis Linnaeus)
C.2 Nisa Rachmania
Mubarik Production and
Application of Inoculant Acid-
Aluminium Tolerant
Bradyrhizobium japonicum
Strains with Soybean
14.00 :14.30 A.3 Purkan
Single Amino Acid Residue
Substitution in two Isolates of
Isoniazid- resistant
Mycobacterium Tuberculosis
Catalase Peroxidase
B.3 Altaf Simiar
Xylanase Production from
Winter Mushroom Flamulina
velutipes grown on optimized
cultural conditions
B.10 Mohammad Sahlan
Mechanism of Interaction
between and Group II
Chaperonin
C.3 Rika Indri Astuti
Detection and Cloning of a
Gene Involved Zwetermicin A
Biosynthesis from Plant
Growth Promoting
Rhizobacterium Bacillus sp
CR64
14.30: 15.00 A.4 Elisa Friska Romasi
Analysis of Alpha -
Glucosidase Inhibitors
Extracted From Several
Medical Herbs
B.4 Diana Lo
The Prospective of Raw
Material and Pathway
Modification in Polylactic
Acid Production as
Ecofriendly Biodegradable
Plastic
B.11 Fernita Puspasari
Cloning of A soft Coral
Associated Bacillus
aquimaris α-Amylase Gene
C.4 Tety Hartatik
Analysis Polymorphism Gene
in Cattle by DNA Sequencing
15.00:15.30 C O F F E E B R E A K
Chairwoman : Dr. Elfahmi Chairwoman : Dr. Ernawati
A.
Chairwoman : Dr.
Maelita Ramdani M.
Chairman : Dr. Adi Pancoro
Biotechnology of Human Health Biotechnology of Industrial and
Application
Biotechnology of Industrial
and Application
Biotechnology of Agricultral
Productions
15.30:16.00 A.5 Ragil Yoga Edi
From Science to farming and
the Seek for the Future Policy
on agricultural Biotechnology
Products Commercializattion
B.5 Akbar Ciptanto
Ethanol Production from
Glycerol of Biodiesel Plant
using Batch and Semi-
continuous Fermentation by
Enterobacter aerogenes
TISTR 1468
B.12 Lanny Hartanti
Effects of Temperature and
pH on the Capability of
Enterobacter aerogenes as
Hg remediator
C.5 Indi Hendraswari
Assessing Growth And
Microsatellite Diversity Of
Timor Mountain Gum
(Eucalyptus Urophylla St.
Blake) Provenances For Tree
Improvement Programs And
The ITB’s Golden Anniversary
International Conference on
Energy and the Environment: Reinvention for Developing Countries
Bandung, 15th-20
th of June 2009
Breeding Assistance
16.00:16.30 A.6 Ketut Gede Muliartha
DNA Methylation of Rar β2
Promotor is Epigenetic
Process on Breast Cancer
B.6 Murdinah
Biodegradable Plastic Made
Hidrocoloid Alginate Extracted
from Sargasum filifend
B.13 Sandhya Babel
Lignin recovery from pulp
and paper mill wastewater
C.6 Priya Chatterjee
Neuroethics: The Pros and Co
16.30:17.00 A.7 Reinhard Pinontoan
Identification of a Calcium-
Dependent Protein Kinase
Gene Involved in
Osmoregulation in
Chlamydomonas reinhardtii
B.7 Mohammad Faseleh
Jahromi
Solid state fermentation for
improvement of fatty acids in
byproducts
B.14 Sendy Adhika Noor
Isolation, purification and
characterization of cellulase
from giant snail (Achatina
fulica) and application to
hydrolyze cellulosic
materials
-
17.00 : 19.00 PREPARATION TO DINNER
The ITB’s Golden Anniversary
International Conference on
Energy and the Environment: Reinvention for Developing Countries
Bandung, 15th-20
th of June 2009
Day 2 : TUESDAY, JUNE 16th 2009
TIME PARALLEL SECTION
VENUE : WEST HALL VENUE : EAST HALL - 1 VENUE : EAST HALL – 2 VENUE : EAST HALL - 3
- Chairwoman : Dr. Marselina
Irasonia Tan
Chairman : Dr. Zeily
Nurachman
Chairman : -
- Biotechnology of Human Health Biotechnology of Industrial and
Application
Biotechnology of Agricultral
Productions
08.30 - 09.00 - A.8 Ricardo Ubyaan
Mitochondrial DNA Variations in
Indigenous Populations of Southern
Papua and its implications on
Native Papuan Haplogroups
B.15 Suraya
The Prospect of Gadung
(Dioscorea hispida Dennst).
Tuber as Carbon Source for
Aspergillus Niger to Produce
Amylase
-
09.00 - 09.30 - A. 9 Uun Yanuhar
The Role, Expression and
Production of Brian Receptor
Protien Of Humpback grouper
Cromileptes altivelis to Viral
Nervous Necrotic Infection for
Development of Diagnostic
Material
B.16 Tania Surya Utami
Nitrous Oxide Gas Removal
Using Compost Biofilter
-
09.30 – 10.00 - A.10 Yohanis Ngili
Nucleotide Polymorphism on D-
Loop Human Mitochondrial DNA
from Endoderm, Mesoderm, and
Ectoderm
- -
10.00 - 10.30 C O F F E E B R E A K
VENUE : EAST HALL
Session PLENARY (Chairman : Dr.Ronny Purwadi )
10.30 - 11.00 Toward Self Sufficient in Food and Biopharmaceutical Through Biotechnology by Dr. Sony
Suhandono (School of Life Sciences and Technology – ITB).
11.00 – 11.30 Marine Biotechnology in Indonesia : Present status and future Development by Prof. Dr.
Hari Eko Irianto (Director, Research Center for Marine and Fisheries product Processing and
Biotechnology, Ministry of Marine Affairs and Fisheries).
11.30 – 12.00 Overview of current status and development strategy of biotechnology in Indonesia by Dr.
Bambang Prasetya
(Head of Biotechnology Consortium Indonesia
12.00 - 13.00 L U N C H B R E A K
13.00 - 13.30 Millennium Research Agenda
FORUM for Biotechnology.
Chairman :
Dr. Adi Pancoro
Dr. Bambang Prasetya
13.30 - 14.00
14.00 - 14.30
14.30 - 15.00
15.00 - 15.30 C O F F E E B R E A K
The ITB’s Golden Anniversary
International Conference on
Energy and the Environment: Reinvention for Developing Countries
Bandung, 15th-20
th of June 2009
15.30 - 16.00 Millennium Research Agenda
FORUM for Biotechnology:
Dr. Adi Pancoro
Dr. Bambang Prasetya
16.00 - 16.30
16.30 - 17.00
17.00 - 19.00 PREPARATION TO DINNER
SEAN
138
Analysis Polymorphism Gene in Cattle by DNA
Sequencing
Tety Hartatik, Tri Satya Mastuti Widi and Muhammad Cahyadi
Department of Animal Production, Faculty of Animal Science,
Gadjah Mada University, Yogyakarta
Jl. Fauna No. 3 Bulaksumur Yogyakarta 55281
ABSTRACT
Studying research technologies on molecular biology is useful to improve the understanding on
molecular genetics in animal. The study of genetic variation of animal population involves the change
of allele size, genotype and phenotype. One of the molecular-based types of genetic variation is the
polymorphism of DNA sequences. Polymorphism can be identified by investigating the changes in
nucleotides which could be transition or tranversion and insertion or deletion. Mitochondrial DNA
(mtDNA) restriction fragment length polymorphism (RFLP) are useful tools for studying genetic
variation and in elaborating relationships between and within species and populations. Here we
examined the polymorphism genetics using Polymerase Chain Reaction (PCR)-base methods, PCR-
RFLP and DNA sequencing for partial mtDNA cyt b gene. We have analyzed the 464 base PCR
product of mtDNA cytochrome b gene from domestic cattle with HaeIII and HinfI restriction enzyme.
The result showed two cutting site of Hae III and one cutting site of HinfI. HaeIII produced three
fragments of 126 bp, 159 bp and 179 bp, but HinfI produced two fragments at 305 bp and 159 bp. In
this study, we also examined partial mtDNA cyt b sequence of local cattle in Indonesia such as
Madura cattle, Ongole hybrid cattle (PO), local cattle of Pacitan and crossing bred of those cattle with
exotic breed (Limousin). Madura cattle was considered as a result of cross breeding between Bos
Javanicus (Bali cattle), and Bos indicus (Ongole cattle). Local cattle of Pacitan have a distinct
phenotype with both Madura cattle and Ongole hybrid cattle, but genetically local cattle of Pacitan
have the same branch with Madura cattle. Thus, based on analysis polymorphism of cytochrome b
gene in cattle shows the diverge branch between species.
Keywords: PCR-RFLP, mtDNA cyt b, Madura cattle, local cattle of Pacitan
INTRODUCTION
Cattle are one of the most economically important domestic animals in the world. Their origin,
genetic diversity, conservation and sustainable utilization have received close attention for long time.
Bali cattle are the original cattle in Indonesia. Bali cattle were domesticated from the Banteng (Bos
Javanicus), a wild cattle species. The other local cattle breed in Indonesia is the Madura cattle. It was
considered as a result of cross breeding between Bos Javanicus, and Bos indicus. Bos indicus (Ongole
cattle) was imported to Indonesia in 1930s. Ongole were used in cross breeding programs to generate
Ongole hybrid (PO) cattle as they can adapt to hot and humid climate and a good type of working.
Madura cattle are similar to Javanese cattle, indigenous hybrids with genes from the Bos indicus, Bos
taurus and Bos (bibos) species (Namikawa,1981 cited by Payne and Hodges, 1997). The local cattle
139
of Pacitan are expected as Javanese cattle which lived in East Java have the phenotype of various
kinds coat color.
To study the genetic variation of domestic cattles in Indonesia, we examined the sequence of
mt-DNA cytochrome b. Mitochondrial DNA evolves much faster than nuclear DNA and thus contains
more sequence diversity compared to nuclear DNA, facilitating the identification of closely related
species (Brown et al., 1996). In addition, maternal inheritance of the mt-DNA generally results in lack
of heterocygosity. Mitochondrial DNA of several vertebrates, including mammals, was mainly
investigated for evolutionary studies (Wolf et al., 1999). Comparisons of nucleotide sequences of the
complete mt-DNA or of single genes, e.g. cytochrome b, were used to study inter- and intraspecies
relationships of animals in order to establish the molecular phylogeny (Birstein and DeSalle, 1998;
Birstein et al., 1998).
MATERIALS AND METHODS
Blood and ear tissue samples of cattle were collected from the smallholder of cattle in Yogyakarta,
Madura and Pacitan. As a control sample, the semen of Limousin were obtained from the inseminator
in those region. DNA was isolated from the whole blood and ear tissue using standard SDS/proteinase
K extraction (Sambrook et al, 1989) and DNA KIT high pure PCR template preparation (ROCHE). A
fragment of the mtDNA cyt b was amplified by PCR methods from 50 ng templates DNA with the
primers L14735 and H15149 (Wolf et al, 1999). PCR product were purified with the QIAquick
isolation system (Qiagen, USA) and were sequenced using BigDye Terminator v3.1 Cycle sequencing
Kit and an ABI PrismTM
310 sequencer Systems (Applied Biosystems). The prediction of restriction
enzyme sites was performed base on sequence results. A phylogenetic tree using partial mtDNA cyt b
sequence was constructed using ClustalW 1.81.
RESULTS AND DISCUSSIONS
The mtDNA cytb gene has been widely considered as one of the preferential DNA targets. In this
study, we analyzed the 464 bp PCR product of mtDNA cytochrome b gene from domestic cattle with
HaeIII and HinfI restriction enzyme. The result of restriction enzyme site analysis showed mostly the
domestic cattle of Indonesia have two cutting site of Hae III and one cutting site of HinfI. Fifty two
samples of 62 analyzed (84%) showed a digested PCR product with HaeIII produce the fragments of
126 bp, 159 bp and 179 bp. Eighty nine persen of the samples was cut with HinfI and produce two
fragments at 305 bp and 159 bp
Base on the mtDNA cyt b sequence alignment analysis, local cattle of Pacitan (Pc13) have similar
sequence similarity to Madura cattle (M1) with score 99%. We found polymorphism of mtDNA
cytochrome b gene. Only four single nucleotides were detected in 400 bp nucleotides sequence. The
four points of difference nucleotide were identified as transversiton (AC→AG), deletion (TT→T- )
and transition (AG→GA) (Figure 1). On the other hand, PO cattle of Pacitan (PO1P) and PO cattle of
DIY (PO68) showed higher number of point’s mutation. There are 34 different nucleotides (Figure 2)
which consist of two tranversions (T→G, T→A), one deletion (TC→T-) and 31 transition
((T→C(13), C→T(11), A→G(4), G→A(3)).
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Figure 1. Multiple Sequence Alignments of partial mtDNA cyt b in Madura Cattle (M1) and Local cattle of
Pacitan (Pc13)
Figure 2. Multiple Sequence Alignments of partial mtDNA cyt b in PO Cattle of DIY (PO68) and PO cattle of
Pacitan (PO1P).
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The exterior phenotype of PO1P and PO68 was similar, however we found 34 nucleotides
differences. On the contrary, Madura cattle (M1) has different coat color with Pc13, but the similarity
of DNA sequences were higher than those of PO1P and PO68. Limpo cattle of Pacitan were derived
from crossing of three breeds, Limousin, PO and local cattle itself. We compared mtDNA cyt b in
Limousin and Limpo cattle of Pacitan at Figure 3. There are 31 different nucleotides which consist of
two tranversions (T→G, C→A), one transition-deletion (TT→C-) and 28 transition ((T→C(11),
C→T(11), A→G(4), G→A(2)).
Sequence analyses indicated that mtDNA cyt b gene for Local cattle of Pacitan were closely
related to Madura cattle. Local cattle of Pacitan has distinct sequences significantly from the Ongole
hybrid (Bos indicus) and Limousin (Bos Taurus) sequences with several point mutation, tranversion,
transition, and deletion/insertion.
mtDNA is a marker of maternal lineage, which in cattle corresponds to the history of the herd
(Nijman et al, 2003). Neighbor-Joining tree using of mtDNA cytochrome b was constructed using
ClustalW 1.81.
Figure 3. Multiple Sequence Alignments of partial mtDNA cyt b in Limpo Cattle of Pacitan and Limmousine
(Lim).
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Figure 4. Neighbor-Joining tree was constructed using ClustalW 1.81. The individual sample represented local
cattle from Pacitan (1,2,3), Madura cattle (4), Limousin as exotic breed (5) and Ongole hybrid (6).
The existence of three groups ((Bos Taurus(5), Bos Indicus (6) and Bos Javanicus (1-4)) was
confirmed (Figure 4). Madura cattle and local cattle of Pacitan shows the diverge branch from Bos
Taurus and Bos Indicus. This branch indicate that local cattle of Pacitan and the new hybrids of these
cattle are originated from Bos Javanicus.
CONCLUSION
Local cattle of Pacitan indicate the spesific characteristic exterior and specific mtDNA cyt b. For
preventing genetics erosion of local cattle by cross breeding, further investigation on local cattle in
Indonesia is necessary. It is needed to study the molecular genetics of local cattle and to conserve the
local genetics resources.
ACKNOWLEDGMENTS
This work was partly supported by Research Grant from Gadjah Mada University through Hibah
Bersaing XVI year 2008. We thank Prof. Masahiko Hatano and Prof. Takeshi Tokuhisa for laboratory
facilities and valuable things related to sequencing DNA. We thank Mrs. Hanasono for technical
guidance in Biomedical Research Center, Chiba University. We also thank the staff of veterinary
services in Pacitan Regency and Madura island, and the students who spent the time for sample
collection.
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