Species specific polymerase chain reaction (PCR) assay for identification of pig (Sus domesticus) skin in “Rambak” crackers - repository civitas UGM

  Jointly organized by

  Volume II PROCEEDINGS st

1 International Conference on

  Tropical Animal Science and Production (TASP 2016)

  “Integrated Approach in Advanced Animal Science and Innovation Technology” July 26-29, 2016

  Ambassador Hotel, Bangkok, Thailand

  

Volume II

Proceedings

st

1 International Conference on

  

Tropical Animal Science and Production

(TASP 2016)

“Integrated Approach in Advanced Animal Science

and

  Innovation Technology”

  July 26-29, 2016 Ambassador Hotel, Thailand

  Jointly organized by:

  Suranaree University of Technology Rajamangala University of Technology Isan Nakhon Ratchasima Rajabhat University Thailand Research Fund

  Welcome Letter from the Rector of SUT Dear Participants,

  I would like to welcome all of you who have come to Bangkok, Thailand, from all over the world. We are proud and honored to host

  st

  the 1 International Conference on Tropical Animal Science and Production (TASP 2016). This year, Suranaree University of

  th

  Technology is celebrating its 26 Anniversary. Over the past 26 years, we are proud to be consistently ranked as one of Thailand’s leading universities, with a strong reputation for research excellence and impact. This Conference is certainly a wonderful addition to our

  th calendar of 26 anniversary celebration activities.

  The theme of this conference “Integrated Approach in Advanced Animal Science and

  Innovation Technology” is appropriate in view of the orientation of technological and institutional changes in such a manner as to ensure the attainment and continued satisfaction of human needs for present and future generations. If you take a look at the conference program, you will agree that this conference is going to be busy and productive. There is a lot to offer for a four-day event. I would like to take this opportunity to thank the Conference Organizing Committee for their diligent work. I would also like to thank participants, especially those of you coming from abroad, for joining us and sharing your valuable experience and ideas. I hope this conference provides you with a forum to exchange scientific ideas, inspire new research, and new contacts for closer co-operation, so that we can, together, envision the future of a promising development of the animal science and production in the Tropic and all over the world.

  Last but not least, I sincere thank our co-hosts, namely, Rajamangala University of Technology Isan (RMUTI), Nakhon Ratchasima Rajabhat University (NRRU) and The Thailand Research Fund (TRF), and our sponsors for their immense support in making this conference successful.

  In closing, I wish you many splendid ideas, new partnerships and the best of success, and I wish our visitors from abroad will have a very pleasant stay in the beautiful city of Bangkok.

  Sincerely, Prof. Dr. Prasart Suebka Rector

  Message from the Chairman of TASP 2016 Dear Participants,

  On behalf of the Organizing Committee, I would like to welcome you to

  st

  The 1 International Conference on Tropical Animal Science and

  th th

  Production 2016 (TASP 2016) in Bangkok, Thailand, from 26 to 29 July 2016. The theme of this conference is “Integrated Approach in Advanced Animal Science and Innovation Technology” to appropriate in view of the orientation of technological and institutional changes in such a manner as to ensure the attainment and continued satisfaction of human needs for present and future generations. The purpose of the conference is to provide an opportunity for animal scientists, animal agriculture-crop scientists and those from related disciplines to discuss and debate research issues relating to aquaculture, animal behavior and welfare, animal genetics and breeding, animal physiology and reproduction, animal biotechnology, epigenetics, feed science and technology, livestock farming system, livestock management, meat science, non-ruminant nutrition, nutrigenomics, pasture, pet, ruminant nutrition, veterinary and others.

  st

  The 1 TASP 2016 is organized by 4 institutes; Suranaree University of Technology (SUT), Nakhon Ratchasima Rajabhat University (NRRU), Rajamangala University of Technology Isan (RMUTI) and The Thailand Research Fund (TRF).

  st

  I sincerely hope that the papers presented in the 1 TASP 2016 would cover all major areas animal science and production and yield meaningful information leading to further strengthening of animal science and production in the tropical areas. I would like to thank all authors for submitting their works and all members of Local and International Organizing Committee, reviewers, all co-host institutions, sponsors without the support of whom a conference of this scale would not have possible, and all individuals who have contributed greatly to the success of this conference.

  Sincerely, Assoc. Prof. Dr. Pramote Paengkoum Chairman, Organizing Committee TASP 2016

  Message from the Chairman of Academic Committee Dear Participants,

  On behalf of Academic Committee, it is my great pleasure to welcome

  st

  you to the 1 International Conference on Tropical Animal Science and Production (TASP 2016) in Bangkok, Thailand. The conference has been designed to provide an opportunity for animal scientists, researchers and research scholars to exchange and to share their experiences, and to promote awareness of the ongoing research achievements. It also provides the premier interdisciplinary forum for animal scientists to present their latest research results, ideas, developments, and applications in all areas of Animal Science and Production. The conference will feature a program of keynote and plenary speakers, invited speaker and oral sessions, posters, and workshops. I would like to express my sincere thanks for all keynote speaker, plenary speakers, invited speakers and all participants for their contribution to make this scientific meeting a great success and fruitfulness. It is our great honour to have Prof. Dr. Charan Chantalakhana from Thailand, Prof. Dr. Metha Wanapat from Thailand, Prof. Dr. Liang Chou Hsia from Taiwan, Assoc. Prof. Dr. Jowaman Khajarern from Thailand, Prof. Dr. Junichi Takahashi from Japan, Assoc. Prof. Dr. Pongchan Na-Lampang from Thailand, Dr. Pascal Mermillod from France, Dr. John Moran from Australia, Prof. Dr. Yang Sheng Lin from China, Prof. Dr. Mongkol Techakumphu from Thailand, Prof. Dr. Thomas J. Schonenwille from the Netherlands and Prof. Dr. Nguyen Van Thu from Vietnam, who are the esteemed speakers and specialists on their own fields to joint us in this conference. I also deepy appreciate the scientific committee, reviewer and editorial boards for their great contribution to make this conference efficacious. I hope that you will enjoy the conference and your interaction with your colleagues from many different countries. Also, I wish that this conference will stimulate a creative exchange of ideas and opportunity for networking, collaboration, sharing of technical information and the building of trust relationship internationally. Last but not least, I am confident that you will enjoy your visit in Bangkok and find a harmonious blend of culture, cuisine, arts, and nice sightseeing along excellent shopping opportunities.

  Yours sincerely, Asst. Prof. Dr. Sutisa Khempaka Chairman of Academic Committee

  List of TASP 2016 Committee International Advisory Committee

  Assoc. Prof. Dr. Pongchan Na-Lampang (Thailand) (Advisor) Assoc. Prof. Dr. Pramote Paengkoum (Thailand) (Chairman) Asst. Prof. Dr. Sutisa Khempaka (Thailand) (Secretary) Prof. Dr. Ermias Kebreab (USA) Prof. Dr. Chris McSweeney (Australia) Assoc. Prof. Dr. Jatuporn Krajaysri (D.V.M.) (Thailand) Prof. Dr. Juan Boo Liang (Malaysia) Asst. Prof. Dr. Pakanit Kupittayanont (D.V.M.) (Thailand) Prof. Dr. Zhongton Yu (USA) Assoc. Prof. Dr. Songsak Chumpawadee (Thailand) Asst. Prof. Dr. Thomas Schonewille (The Netherlands) Asst. Prof. Dr. Chalermpon Yuangklang (Thailand) Prof. Dr. Yasuhiro Kawamoto (Japan) Asst. Prof. Dr. Pipat Lounglawan (Thailand) Prof. Dr. Pietro Celi (USA) Assoc. Prof. Dr. Surintorn Boonanuntanasarn (Thailand) Prof. Dr. Nguyen Van Thu (Vietnam) Assoc. Prof. Dr. Anut Chantiratikul (Thailand) Prof. Dr. Hsia Liang Chou (Taiwan) Dr. Samorn Ponchunchoovong (Thailand) Dr. Viengsakoun Napasirth (Laos) Dr. Wittawat Molee (Thailand) Prof. Dr. Long Ruijun (China) Asst. Prof. Dr. Amornrat Molee (Thailand) Organizing Committee of TASP 2016 Advisors Assoc. Prof. Dr. Pongchan Na-Lampang Assoc. Prof. Dr. Wisitiporn Suksombat Chairman Assoc. Prof. Dr. Pramote Paengkoum Vice Chairmen Assoc. Prof. Dr. Jatuporn Krajaysri Asst. Prof. Dr. Chalermpon Yuangklang Secretariat Assoc. Prof. Dr. Surintorn Boonanuntanasarn Scientific Committee Asst. Prof. Dr. Sutisa Khempaka Asst. Prof. Dr. Pipat Lounglawan

  

TASP 2016. Integrated Approach in Advanced Animal Science and Innovation

Technology: Proceedings. Volume II: 343 pages.

  First Edition: 200 books

  ISBN: 978-974-533-710-7 Year 2016

LERDSILP PRINTING

  336 Suranaree Sub-district, Muang Nakhon Ratchasima District, Nakhon Ratchasima 30000, Thailand

  The individual contributions in this publication and any liabilities arising from them remain the responsibility of the authors. The publisher is not responsible for possible damages, which could be a result of content derived from this publication.

  

Contents

  Welcome Letter from the Rector of SUT a

  Message from the Chairman of TASP 2016 b

  Message from the Chairman of Academic Committee c List of TASP 2016 Committee d

  Animal Behavior and Welfare

  FO0065 Noncage housing systems and laying hen welfare: keel bone damage

  2 and feather pecking

  M. Petek

  FP0422 Application of geo informatics for wild elephant habitat in Thap Lan

  6 National Park

A. Srisuwan, and W. Panurak

  Animal Biotechnology

  FO0036 The effect of using Solanum dubium and vinegar as alternatives to

  12 rennet enzyme on sensory characteristics of the white cheese (Gibna

  bayda

  )

  Ibrahim Elimam, Mohammed Elawad, and Elgazafey Basheer

  FO0282 Species specific polymerase chain reaction (PCR) assay for

  16 identification of pig (Sus domesticus ) skin in “Rambak” crackers

  

Y. Erwanto, R. Yuliatmo, Sugiyono, A. Rohman, and Sismindari

  FP0262 Chromosomal analysis to predict early fertility in bovine bulls

  22 M. F. Saad, A. M. Saeed, and H. A. El-naggar

  Animal Genetics and Breeding

  FO0016 Defining economic values of important traits in one hump camel in

  27 desert areas rearing system

  M. Vatankhah

  FO0072 Cow efficiency in crossbreeding systems as defined by kilogram calf

  31 weaned per large stock unit

  M. C. Mokolobate, M. M. Scholtz, F. W. C Neser, and A. Theunissen

  FO0092 Association between STAT1 gene polymorphism and milk production

  35 traits in Holstein, Jersey and Turkish native cattle breeds

  

O. Cobanoglu, E. K. Gurcan, S. Cankaya, E. Kul, and H. S. Abaci

  FO0162 Changes in the cow productivity of the indigenous Afrikaner breed and

  39 its environmental impact

  

M. M. Scholtz, F. J. Jordaan, M. C. Mokolobate, A. Maiwashe, Z.

King, and F. W.C. Neser

  FO0244 Production performance of red jungle fowl versus native roosters

  43 under confinement system: a comparative study

  Francisco F. Buctot JR. FO0308 Polymorphisms of GH|EcoRV and GHR|Eco72I loci in Indonesian

  49 indigenous chicken

  Isyana Khaerunnisa, Melati Ardiyana, Candra Krisdianto, Sri Darwati, Tike Sartika, and Cece Sumantri Animal Physiology and Reproduction

  FO0084 Influences of location and ecological zones on fresh cow milk

  55 composition from milk vendors in Nigeria

  D. Zahraddeen, D. Y. SalihuH. O. Okin-Aminu, and A. A. Mohammed

  FO0129 Characterization and structure of the mammary gland of rabbit related

  59 to milk production and prolactin blood levels recorded during lactating period

  T. T. Amroun, N. Zerrouki-Daoudi, and M. Charlier

  FO0152 Reliability of placentome diameter for determination of gestation using

  64 transabdominal ultrasonography in sheep and goats

  M. A. Waziri, M. M. Bukar, A. Alapa, and A. Y. Ribadu

  FO0241 Pregnancy and full-term development of embryos derived from natural

  68 mating and AI following estrous synchronization in swamp buffalo

  B. Rosadi, T. Sumarsono, and D. Darmawan Aquaculture

  FP0179 Potential aquaponic culture in the Mekong Delta

  72 Vu Ngoc Ut, Nguyen Quoc Linh, and Huynh Truong Giang FP0188 Effect of supplementation of attractants to diets on feeding behavior

  76 and growth performance of white shrimp (Litopenaeus vannamei)

  O. Jintasataporn, J. Matas Ballarin, A. Hung, and M. L. He

  FP0395 The effect of replacement of fishmeal by rice wine residual on growth

  80 performance and intestinal morphology of Thai Panga

  K. Taveewannaboon, and S. Ponchunchoovong

  FP0420 Growth of Thai Climbing Perch (Anabas testudineus) and snakeskin

  85 gourami (Trichogaster pectoralis) in brackish water from saline soil distribution impact area in Nakhon Ratchasima province

  N. Noinumsai, S. Wongma, and P. Ngoaknaiwang

  FP0428 Microencapsulation of Saccharomyces cerevisiae and its effects on

  94 fish intestinal microbiota

  K. Ditthab, S. Boonanuntanasarn, and S. Chaijamrus Feed Science and Technology

  FO0117 Effect of beneficial microorganisms supplement diets on growth

  99 performance of post weaned pigs in the Mekong Delta, Vietnam

  L. T. Men, N .T. D. Trang, H. M. Tri, V. V. Son, and N. D. Hien

  FO0320 The feed digestibility of quails as affected by administration of 104 encapsulated probiotic product

  U. Kalsum, F. Wadjdi, and L. Rahardjo

  FO0360 Treatment of high-tannin madre de cacao (Gliricidia sepium) forage 108 and crude protein utilization in native goats (Capra hircus)

  Hershey P. Mondejar, and Lolito C. Bestil FO0383 Feed intake, digestibility, growth performances and eating behavior of 112 Thai native beef cattle fed diets differing in energy density using cassava pulp with rice straw

  T. Gunha, K. Kongphitee, and K Sommart

  FO0386 Ensilage quality, digestibility and enteric methane emission of the 116 fermented total mixed ration in Thai native beef cattle

  K. Kongphitee, and K. Sommart

  FP0013 Effect of the incorporation of brewers' grains on growth performance 121 of broilers

  F. Arbouche, R. Arbouche, H. S. Arbouche, and Y. Arbouche

  FP0193 Effect of functional palatability enhancers with milk cheese flavors on 125 feed intake and growth performance of weanling piglets

  H. F. Wang, X. D. Wang, X. X. Dou, L. Liu, J. L.Wang, and M. L. He

  FP0197 Effect of functional palatability enhancers with colostrum flavors on 129 feed intake and growth performance of weanling piglets

  F. Li, S. Wu, Y. Zhang, and M. L. He

  FP0359 Effect of catfish by-products on growth performance of growing 133 Muscovy ducks

  T. L. Nguyen, and T. K. D. Nguyen Livestock Farming System / Pasture

  FO0103 Potential benefits to dairy industry productivity through breeding more 138 digestible tropical grasses utilizing contemporary approaches

  B. C. Granzin

  FO0364 The addition of biogas sludge and cattle waste compost towards 142 soybean productivity through the use of saturated soil culture (SSC) technology on tidal swamp

  S. A. A. Tani, B. P. Purwanto, M. Ghulahmadi, A. M. Fuah, Salundik, and W. A. Ridwan

  FO0384 Husbandry systems of keeping native chickens in tropical areas of 147 Indonesia

  Yusuf L. Henuk, and Darma Bakti

  FP0014 Effects of phenological stage of flood meadows Northeastern Algeria 151 on their production and nutritional values

  R. Arbouche, Y. Arbouche, H. S. Arbouche, and F. Arbouche

  FP0015 Effects of phenological of Algerian Northeastern forest permanent 155 grassland on their production and nutritional value

  H. S. Arbouche, R. Arbouche, Y. Arbouche, and F. Arbouche

  FP0234 Elements that define a livestock system intensive silvopastoral in 160 Mexico

  

G. Salas-Razo, D. Ramírez-Martínez, M. Perea-Peña, E. Gutiérrez-

Vázquez, R. Garcidueñas-Piña, and J. Espinoza-Villavicencio

  FP0367 Analysis of technologies used in the livestock system intensive 164 silvopastoral in Mexico and the technological changes that could improve efficiency

  

G. Salas-Razo, D. Ramírez-Martínez, M. Perea-Peña, E. Gutiérrez- Vázquez, R. Garcidueñas-Piña, and J. Espinoza-Villavicencio FP0423 Effects of plant spacing on yields of Napier Pak Chong1 during dry 168 season (Pennisetum purpureum x Pennisetum americanum)

  

S. Panatung, S. Boonsari, A. Suntornpithug, and C. Charoenrporn

Livestock Management

  FO0083 Evaporative cooling has potential to improve performance of Zebu 173 cattle in tropical environments

  Olusola A. Olorunnisomo

  FO0093 Impediments, interventions and impact assessment for dairy farmers in 177 Pakistan: A roadmap for improved rural livelihoods

  Z. Mustafa, and M. N. M. Ibrahim

  FO0094 Effect of water and balanced diet on milk production of dairy animals 181 in Pakistan

  M. S. Ashraf, Z. Manzoor, N. Khan, T. Abbas, Z. Mustafa, and M. N. M. Ibrahim

  FO0101 Small ruminants supplemental feeding regimes in arid agro-pastoral 185 systems in Pakistan

  A. Razzaq, M. Islam, B. Ann Rischkowsky, M. Louhaichi, and M. N. M. Ibrahim

  FO0219 A framework on vulnerability analyses of Southeast Asian countries 189 on goat husbandry to climate change and food security

  S. Ocak, B. Gürer, and O. B. Karaca

  FP0109 Analysis of productivity factors according to the feeding levels of 194 automatic milking system installed dairy farms

  I. S. Nam, Y. S. Yang, B. M. Heo, W. Y. Kim, J. M. Yeo, K. T. Nam, and K. M. Chang Meat Science

  FO0033 The pH of kid meat influence by diet nutrients 198

  M. Karami

  FP0216 Effect of white sesame seeds and cayenne pepper on quality in reduced 201 sodium and low-fat precooked pork patties

  Jen-Hua Cheng

  FP0363 Variations in meat chemical composition and quality characteristics of 205 different Saudi sheep breeds

  G. M. Suliman, A. M. El-Waziry, A. N. Al-Owaimer, E.O.S. Hussein,

  H. Metwally, and M. S. Al-Harbi Non Ruminant Nutrition

  FO0077 Effect of protein and energy levels on performance and egg quality of 209 Black-boned layers (Ac chicken) at early first production cycle

  

Truong Van Phuoc, Nguyen Nhut Xuan Dung, and Luu Huu Manh

  FO0078 Effect of dietary rice bran oil combination with salmon fish oil on 214 reproduction performance, fatty acid content of yolk in layers

  

Le Thanh Phuong, Luu Huu Manh, and Nguyen Nhut Xuan Dung FO0145 Effect of dietary crude protein levels on nutrient intakes, growth rate 219 and economic return of Guinea fowls in Mekong delta, Vietnam

  T. K. D. Nguyen and V. T. Nguyen

  FO0265 Effect of dietary supplementation with green tea powder on 224 performance characteristic, meat organoleptic quality and cholesterol content of broilers

  

Nguyen Hoang Thinh, Nguyen Thi Vinh, Bui Huu Doan, and Pham

Kim Dang

  FO0322 Performance of layers fed diets supplemented with methionine 229 producing probiotic microorganisms

  

B. N. Suresh, M. C. Shivakumar, Nagaraj Naik, N. Suma, and L.

Manjunatha

  FO0327 233

  Performance and production of functional duck’s egg fed diets containing Indigofera zollingeriana leaves meal and Lemuru fish oil

  Sumiati, A. Darmawan, and W. Hermana

  FO0361 Effects of the mixture of essential oils and organic acids on 238 performance, carcass yield and internal organs in broilers

  Sakine YALÇIN, Handan ESER, Suzan YALÇIN, and İlyas ONBAŞILAR

  FP0187 Comparison on effects of two palatability enhancers on growth 241 performance, intestinal villi and relative taste receptor gene expression of piglets

  T. Huang, C. Xia, J. Zuo, Z. Wang, and M. L. He

  FP0311 Effect of curcumin powder on productive performance and 245 digestibility of turkeys

  Kraisit Vasupen, Chalermpon Yuangklang, Sasiphan Wongsuthavas, Benya Saenmahayak, and Smerjai Bureenok

  FP0338 The effects of methionine and its relationship to tryptophan on feed 248 intake and growth performance of weaning piglets

  W. Jantasin, and L. C. Hsia

  FP0419 Effects of supplementation of Pro L. in water on production 253 performance in broiler

  P. Sutatta, U. Laedthaisong, and S. Paengkoum

  FP0425 Reproductive performance and fertility response of laying hens as 256 affected by dietary energy and antioxidant substance supplementation

  

P. Pasri, S. Khempaka, W. Molee, A. Molee, N. Gerard, and P.

Mermillod

  Ruminant Nutrition

  FO0128 The effect of concentrate containing a mixture microbes on in vitro 261 nutrients digestibility and fermentation characteristics of King grass

  B. Santoso, T. W. Widayati, B. Tj. Hariadi, and H. Abubakar

  FO0230 Solid state fermentation of wheat straw on large scale for livestock 266 feed in Pakistan

  S. Faisal, A. Muhammad, and A. S. Chudhry FO0240 Effect of the level of palm oil decanter meal preserved with cinnamon 270 bark powder in diet on the rumen environment of cross breed Ettawa goat

  M. Afdal, Jul Andayani, and Hajar Setyaji

  FO0329 Feed additive of betel leaves meal (Piper betle L.) use on ruminants as 273 one of methane mitigation efforts

  A. Sudarman, I. Y. Marcelina, and A. Jayanegara

  FO0343 Assessing the potential of oil palm frond juice as animal feed supplements 277 by determining its nutrients, lignocellulosic and sugar contents

  

N. D. Rusli, K. Mat, C. H. Hasnita, M. Wan Zahari, K. Azhar, M.

Zamri-Saad, and H. A. Hassim

  FO0412 Body weight gain and feed efficiency of young thin-tailed sheep raised 283 under intensive feeding at different level of protein

  

Ari Prima, Nadlirotun Luthfi, Edy Rianto, and Agung Purnomoadi

  FP0074 Feedlot performance of South African Nguni cattle fed low and high 287 energy rations

  D. A. Linde, M. M. Scholtz, A. Theunissen, and E. van Marle-Köster

  FP0254 Differences on biological mechanisms related to residual feed intake in 291 dairy cows

  Y. M. Xi, D. Q. Zhao, L. Li, Z. Y. Han, and G. L. Wang

  FP0348 Oil enrich omega-3 fatty acid supplementation in Brahman crossbred 296 fattening steers diets on blood parameter

  P. Noosen, P. Lounglawan, and R. Mirattanaphrai Veterinary

  FO0086 Study of the changes of serum Ab titer against Newcastle disease virus 301 at various weeks post vaccination in native chickens

  N. Ghaleh Golab Behbahan, and F. Tavan

  FO0099 Epidemiology, comparative diagnostic options and therapeutic studies 306 against nematodes infecting range sheep and goats in Pakistan

  

A. Razzaq, M. Islam, B. Ann Rischkowsky, and M. N. M. Ibrahim

  FO0121 Identification of major health problems in a breeder goat farm in 310 Sabah, Malaysia

  W. Nieccorita, M. Zamri-Saad, K. Azhar, A. B. M. Zuki, A. H. Hasliza, and A. Punimin

  FO0156 Evaluation and use of 2 cow-side tests for on-site detection of ketone 313 bodies in late gestation does

  A. A. Syahirah, H. A. Hassim, A. A. Saharee, S. S. Syed-Hussain, A. F. M. Azmi, M. Ajat, and R. Mansor

  FO0357 Investigating the seroprevalence of Ornithobacterium rhinotracheale 317 infection in broiler in Fars province, Iran

  

Mohammadjavad Mehrabanpour, and Maryam Ranjabar Bushehri

  Others

  FO0355 Performance, carcass characteristics and meat composition of broilers 322 in response to early feed restriction

  E. E. Onbaşılar, S. Yalçın, E. Torlak, and P. Özdemir

  FO0356 Effects of rearing method in the first week on the performance and 325 meat quality of male and female Pekin ducks

  E. Erdem, E. E. Onbaşılar, A. Kocakaya, and S. Yalçın

  FO0362 Red meat consumption of children aged 12-23 months in Turkey 327

  S. Songül Yalçın, Eda Köksal, Gülden Pekcan, and Başak Tezel

  FP0276 Amylase, protease and xylanase production from Aspergillus niger by 331 solid state fermentation using cassava pulp and corncob as substrat

  Kunthika, V., P. Pianpumepong, and S.Wongsutthawas

  FP0340 Changes in composition of awassi and saanen colostrum during 335 postpartum

  Oya Berkay Karaca, and Sezen Ocak

  FP0417 Phenolic content and antioxidant activities in sunflower sprout 340

V. Tanamool, S. A. Kohnhorst, and A. Chatkon

  st

1 International Conference on Tropical Animal Science and Production (TASP 2016)

  July 26-29, 2016 Ambassador Hotel, Thailand

• - FO0282 -

  Species specific polymerase chain reaction (PCR) assay for identification of pig (Sus domesticus) skin in “Rambak” crackers

• 1,3

  1 2 3,4

  4 Y , R , Sugiyono , A , and Sismindari

  . Erwanto . Yuliatmo . Rohman

  1

  2 Faculty of Animal Sciences, Gadjah Mada University, Yogyakarta, 55281, Indonesia; Faculty

  3

  of Veterinary Medicine, Gadjah Mada University, Yogyakarta, 55281 Indonesia; Research

  4 Center of Halal Products, Gadjah Mada University, Yogyakarta, 55221, Indonesia; Department

  of Pharmaceutical Chemistry, Faculty of Pharmacy, Gadjah Mada University, Yogyakarta, 55221, Indonesia

  Abstract

  “Rambak” is the animal skin crackers product which one of the popular food products derived from the animal by-products in Indonesia . For the religion reasoning the presence of pig derivatives in any food products is prohibited for Moslem community . For this reason, the analytical methods offering accurate and sensitive results are highly needed in order to assure the halal authenticity of food . The objectives of this research is to developed the pig species specific primer for identification of specific pig deoxyribonucleic acid

  (DNA) in “Rambak” cracker using polymerase chain reaction ( PCR) analytical methods. This research used were four kinds of animal skin crackers in individual or mixture samples . Skin mixture were prepared and divided into nine formulas as follows 1 ) 100% pig; 2) pig 90%: cattle 10%; 3) pig 75%: cattle 25%; 4) pig 50%: cattle 50 %; 5) pig 25%: cattle 75%; 6) pig 10%: cattle 90%; 7) pig 1%: cattle 99%; 8) pig 0.1%: cattle 99,9

  %; and 9) 100% cattle. The same formula also prepared in mixture between pig skin with buffalo and goat skin . DNA were isolated from the cracker products and applied in PCR using perimer specific . The optimized PCR assay was subsequently validated for its specificity with deoxyribonucleic acid (DNA) extracted from cattle, buffalo, goat and pig in individual or mixture samples . Results showed the primers designed generated specific fragment of 510 bp length for pig cytochrome b DNA

  . The specificity of the primers was tested on four animal species including pig, cattle, buffalo and goat species . Analysis of experimental mixture meat demonstrated that 0.1% of pig tissues could be detected using specific primer . The specificity of pig-specific PCR provides a valuable tool for identification of pig skin and to avoid its fraudulent substitution and adulteration .

  Keywords : pig species, identification, primer specific, cytochrome b gen

• Corresponding author : erwantougm@gmail.com

  

Introduction

  “Krupuk Rambak” or Rambak cracker, is one of the traditional popular food in Indonesia, made from thick skin of domestic animal . This food is traditionally hand made with traditional process

  . One of the interesting issues in food industry is the authenticity of food products. The quality assurance of food product can involve the alteration of the true labeling of food ingredients,

  

[16] st

1 International Conference on Tropical Animal Science and Production (TASP 2016)

  July 26-29, 2016 Ambassador Hotel, Thailand

  in which the high value of raw materials are substituted by cheaper materials with similar biochemical properties . Cattle or buffalo hide are expensive and can be altered with cheaper skin such as pig skin

  . The presence of pig derivatives such as pig skin, lard, pork and porcine gelatin in any food products is a serious matter for moslem community (Mursyidi, 2013 ), and also for Judaism and

  Hinduism because some forbid their followers to consume any foods containing porcine and its derivatives (Regenstein et al., 2003 ). Therefore, it is necessary to evaluate the presence of these pig derivatives in any products using some instrumental techniques

  . Some previous techniques used are differential scanning calorimetry (Nurrulhidayah et al., 2015; Marikkar et al., 2012) , Fourier transform infrared (FTIR) spectroscopy (Rohman et al., 2011), gas chromatography-mass spectrometry (Nizar et al., 2013 ) , high pressure liquid chromatography ( Saeed et al., 1989; Marikkar et al., 2005), electronic nose (Nurjuliana et al., 2011; Mansor et al., 2011), and proton nuclear magnetic resonance

  (Fang et al., 2013). Some of these methods are too laborious and time consuming, consequently, an analytical technique offering rapid and reliable must be used . One of the promising methods suitable for routine analysis is polymerase chain reaction - restriction fragment lenght polymorphism (PCR-RFLP).

  In the previous research we already developed the new technique based on the PCR

• RFLP technique for meat products determination using cytochrome b gen using universal primer and resulted 360 bp DNA fragment . The determination was continued with restriction fragment length polymorphism using BseD I restriction enzyme and able to cleave amplicon of cytochrome b gen into 221 bp and 138 bp (Erwanto et al., 2012). This technique has been applied in detection of pig derivative contamination in commercial meatball of Indonesia local market and showed positive pig meat contamination of nine from thirty
• nine samples ( Erwanto et al., 2014) . PCR- RFLP technique has been established to determine of meat species contamination but still time consuming and need two stage methods, first the amplification of gen target with universal primer and the second, cleave PCR products using restriction enzyme . Therefore, in this study we developed the species specific primer for distinguish of pig derivative particle based on cytochrome b gen in food derivative products from animal skin .

  

Materials and Methods

Sample preparation and DNA extraction

  Individual skin crackers samples of pig, buffalo, cattle and goat were obtained from local market in Yogyakarta, Indonesia . Samples was prepared in laboratory with separate equipment to prevent cross contamination

  . DNA was extracted and purified from skin cracker samples using Sambrook et al. (1989 ) methods with slight modification. Approximately 30 mg of skins were blended using a commercial blender and placed in a 1 .5 mL microcentrifuge tube. A-500 μL of Tris

• EDTA-NaCl (TEN) buffer, 10 μL Proteinase K and 50 μL 10% Sodium Dodecyl Sulfate (SDS) were added and mixed by vortexing and was incubated at 42°C in a water bath overnight .

  After overnight incubation, 50 μL5 M NaCl, 400 μL phenol and 400 μL chloroform- isoamyl- alcohol was added to the mixture and then incubated at 37°C for 1 h and centrifuged at 3,000 rpm for 5 min . The supernatant was taken and put in a new microcentrifuge tube and mixed by vortexing for 30 s, then 50 μL 5 M NaCl and 1 mL absolute ethanol were added, mixed vigorously and incubated at

• –20°C for 1 h. The DNA was precipitated from the mixture after centrifuging at 8,000 rpm for 5 min, 4°C and the supernatant discarded

  . The 70% ethanol was gently poured off then resuspended DNA for a second precipitation . The solution was then placed at room temperature overnight . The resulting pellet was added with 50 mL Tris-EDTA (TE) buffer and stored at –20°C

  

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  for further analysis . Skin cracker samples were prepared by mixing of pig in buffalo, pig in cattle, and pig in goat at a final concentration of pig at 0; 0

  .1; 1; 10; 25; 50; 75; 90 and 100%.

  Design of pig specific primer

  The nucleotide sequences of mitochondrial Cyt b genes from sus scrofa (pig) stored in Genbank were taken and aligned using Primer3 website . The set of primers used for amplification consisted of Cyt b-FW and Cyt b

• REV oligonucleotides as follows:

  Cyt b FW 5 ’-CCC AGC CCC CTC AAA CAT CTC A-3’, Cyt b REV 5 ’-ATG TAC GGC TGC GAG GGC GGT-3’. Amplification of the Cyt b gene was performed in a final volume of 25 μL containing 100 ng of extracted DNA, 20µL DreamTaq Green PCR master Mix

  (Thermo Fisher Scientific) and 20 µmol of each primer . Amplification was performed with a thermal cycler according to the following PCR step -cycle program: Pre-denaturation at 94°C for 2 min to completely denature the DNA template, followed by 40 cycles of denaturation at 95°C for 36 s, annealing at 53°C for 73 s . and extension at 72°C for 84 s

  . Final extension at 72°C for 3 min followed the final cycle for complete synthesis of elongated DNA molecules . Six microliters of PCR products were mixed with 1 μL DNA Loading Dye (Thermo Fisher Scientific), then were electrophoresed at constant voltage

  (100 V) on 2% agarose gel (TopVision, Thermo Fisher Scientific) for about an hour in 1×TBE buffer, pH 8 .0 and stained by ethidium bromide. A-100 bp DNA ladder (GeneRuller, Thermo Fisher Scientific ) was used as size reference.

  

Results and Discussion

  Total genom DNA form the various animal skin species was isolated using Sambrook et al . (1989) method with slight modification. During processing proteins are denaturalised due to the time and temperatures of process and DNA should be employed for species identification purposes

  . Therefore the susceptibility of the determination of species in food products depend on the susceptibility in DNA extraction, without the extracted DNA the measurement of the detection could not be continued .

  DNA genom of four species could be successfully isolated (data not shown) although there were some smear in goat skin sample . The quality of DNA and the level content of pig in the crackers mix samples is important data for verifying

   the sensitivity of the method.

  To identify pig skin in a large number of both processed and unprocessed skin crackers, we carried out specific pig PCR amplification . With this fragment, pig has been detected in mixtures treated with cattle, buffalo and goat skin crackers. Some commercial products were analyzed which normally have cattle, buffalo or pig in their crackers product including in fresh skin crackers or various processed skin crackers

  . Using the specific PCR amplification, we detected pig in various concentration . Pig was detected until 0.1% contamination in mixture with cattle (Figure 1), buffalo

  (Figure 2) or goat (Figure 3). In our some previous research showed the PCR-RFLP able to detect pig DNA in heated meat mixtures at levels less than 1 % based on fragment of the cytochrome b DNA

  (Erwanto et al., 2012) and Matsunaga et al. (1999) detected 250 pg of beef DNA, in DNA mixtures . In raw pork and beef mixtures, Calvo et al. (2001) detected up to 0.01% beef in pork.

  The choice of the technique to be applied is based on the results of sequence alignments. Specific PCR seems to be best as a ‘ routine’ test, because it is easy, rapid and allows the discrimination of several species at the same time (multiplex PCR), as well as the analysis of mixed

  

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  matrices . It is considered to be a robust method, mainly in comparison with other methods based on the analysis of single nucleotide polymorphisms (Bottero and Dalmaso, 2011).

  Figure 1 . PCR products of cyt b specific gene from different skin product separated by 2% high- resolution agarose gel electrophoresis . M: Marker; 1: 100% pig skin; 2: pig 90%: buffalo 10%; 3: pig 75

  %: buffalo 25%; 4: pig 50%: 50% buffalo; 5: pig 25%: 75% skin; 6: pig 10%: buffalo 90%;

  7 : pig 1%: 99% buffalo; and 8: pig 0.1% buffalo 99.9%; 9: 100% buffalo skin.

  Figure 2 . PCR products of cyt b specific gene from different skin product separated by 2% high- resolution agarose gel electrophoresis . M: Marker; 1: 100% pig skin; 2: pig 90%: cattle 10%; 3: pig 75

  %: cattle 25%; 4: pig 50%: 50% cattle; 5: pig 25%: 75% cattle; 6: pig 10%: cattle 90%; 7: pig 1 %: 99% cattle; and 8: pig 0.1% cattle 99.9%; 9: 100% cattle skin.

  Figure 3 . PCR products of cyt b specific gene from different skin product separated by 2% high- resolution agarose gel electrophoresis . M: Marker; 1: 100% pig skin; 2: pig 90%: goat 10%; 3: pig

  

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  ., J.Y. Goh, M. Tay, H. F. Lau, and S. F. Y. Li. 2013. Characterization of oils and fats by

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References

  International Conference on Tropical Animal Science and Production (TASP 2016) ^st

  This research was funding by project grant from the directorate of higher education, Ministry of Higher Education, Research and Technology, with contract No . 159/LPPM/2015.

  

Acknowledgment

  quantitative method since this conventional PCR analysis is only qualitative.

  .1 % pig skin). However, further research would be needed in order to develop a

  of pig contamination, due to its simplicity, specificity, and sensitivity (with 35 amplification cycles we can detect 0

   is a powerful technique for the identification

  Specific PCR amplification of this fragment

  

Conclusion

  %: 99% goat; and 8: pig skin 0.1% goat skin 99.9; 9: 100% cattle skin.

  75 %: goat 25%; 4: pig 50%: 50% goat; 5: pig 25%: 75% goat; 6: pig 10%: goat skin 90%; 7: pig skin 1

  

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• - Restriction Fragment Length Polymorphism for halal

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  and mutton fat by GCMS and EA

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  .F., S. R. Arieff, A. Rohman, I. Amin, M. Shuhaimi, A. Khatib. 2015. Detection of butter adulteration with lard using differential scanning calorimetry . International Food Research Journal, 22, 832 -839.

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