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J. Trop. Resour. Sustain. Sci. 5 (2017) 117-120

DNA Extraction and Library Preparation Optimization for Metagenomic Analysis of Giant Panda in Malaysia

Suraj Arvind V Rajasegar1, Azrinaaini Md Yusof2, Suganthi Appalasamy1, Seri Intan Mokhtar3, Mohd Tajuddin Abdullah4, Mohd Noor Mat Isa5, Mat Naim Bin Hj Ramli6, Ten Dennis Choon Yung7, Abdul Kadir bin Abu Hashim8, Dengsheng Li9, ChaiWu Li10, Jayaraj Vijaya Kumaran11*

1Fakulti Sains Bumi, Universiti Malaysia Kelantan, UMK Kampus Jeli, 17600 Jeli, Kelantan, Malaysia
2Fakulti Asas Tani, Universiti Malaysia Kelantan, UMK Kampus Jeli, 17600 Jeli, Kelantan, Malaysia
3Institute of Food Security and Sustainable Agriculture (IFSSA), Universiti Malaysia Kelantan, UMK Kampus Jeli, 17600 Jeli, Kelantan, Malaysia
4Institut Penyelidikan Kenyir (IPK), Universiti Malaysia Terengganu, 21030 Kuala Nerus, , Terengganu., Malaysia
5Malaysia Genome Institute (MGI), National Institute of Biotechnology Malaysia (NIBM), Jalan Bangi, 43000 Kajang, Selangor Darul Ehsan, Malaysia
6Zoo Negara, Zoo Negara, Hulu Kelang, 68000 Ampang, Selangor Darul Ehsan, Malaysia
7Jabatan Perlindungan Hidupan Liar dan Taman Negara (PERHILITAN) Semenanjung Malaysia , Jabatan Perlindungan Hidupan Liar dan Taman Negara (PERHILITAN) Semenanjung Malaysia, Km 10, Jalan Cheras, 56100 , Kuala Lumpur, Malaysia
8Jabatan Perlindungan Hidupan Liar dan Taman Negara (PERHILITAN) Semenanjung Malaysia, , Jabatan Perlindungan Hidupan Liar dan Taman Negara (PERHILITAN) Semenanjung Malaysia, , Km 10, Jalan Cheras, 56100 , Kuala Lumpur, Malaysia
98China Conservation and Research Centre of the Giant Panda (CCRCGP), 8China Conservation and Research Centre of the Giant Panda (CCRCGP), Ya’an, Ya’an, China
10China Conservation and Research Centre of the Giant Panda (CCRCGP), China Conservation and Research Centre of the Giant Panda (CCRCGP) , Ya’an, Ya’an, China
11Faculty of Earth Science, Universiti Malaysia Kelantan, Jeli, Kelantan, Malaysia

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Abstract : The objective of this study was to determine the expected minimum number of sequence reads needed to achieve full coverage of the microbial species found in the gut of giant panda This was done by first, analyzing five different mammalian metagenomes namely horse, coyote, whitetail deer, humpback whale and the bottlenose dolphin as metagenomic references; based on an approximate of 1,000,000 sequence read estimation. After rarefaction analysis using MG-RAST Version 3.0 analysis pipeline, an average value of 775,075 reads was found to be sufficient for metagenomic analysis. Next, the fecal matter of giant panda was sampled at Zoo Negara, and DNA extraction was performed. DNA extraction was then subjected to DNA qualification and quantification analysis; where the results show that the samples are still viable and sufficient in yield to be used for library preparation. After library preparation, the samples were sequenced using Illumina™ MiSeq® next generation sequencer. The results of this research serve as a foundation for further studies of the giant panda metagenome.

Keywords : Giant Panda,Illuminaâ„¢ MiSeq,metagenomic analysis