Introduction The Greater Short Nosed Fruit Bat, Cynopterus sphinx (Vahl, 1797) is a species of bat that can be found in Peninsular Malaysia. It is an important seed disperser and it feeds on various fruits and plant parts. This mitochondrial genome is part of an on-going study on the metagenomics of fruit bats in SEA. NADH dehydrogenase subunit 1 NADH dehydrogenase subunit 2 NADH dehydrogenase subunit 3 NADH dehydrogenase subunit 4L NADH dehydrogenase subunit 4 NADH dehydrogenase subunit 5 NADH dehydrogenase subunit 6 1 2 3 4,5,6 7,8,9,10,11 12,13 14 15 17,18,19 16 20 12S 16S Cytochrome b Cytochrome Oxidase 1 Cytochrome Oxidase 111 Cytochrome Oxidase 11 ATP6 ATP8 The Mitochondrial Genome of Cynopterus sphinx Jayaraj Vijaya Kumaran 1,2,* , Mohd Noor Mohd Isa 3 , Ahmad Sofiman Othman 2 , Shahrul Anuar Mohd Sah 2 , Seri Intan Mokhtar 4 1 Faculty of Earth Science, Universiti Malaysia Kelantan, 2 School of Biological Science, Universiti Sains Malaysia, 3 Malaysia Genome Institute, 4 Faculty of Agro Based Industry, Universiti Malaysia Kelantan. Abstract:- The complete mitochondrial genome of Cynopterus sphinx, a widespread and ecologically important fruit bat is first sequenced here using illumina Hiseq 2000 data. The genome was 16848 bps in length and there were 22 transfer RNA (tRNA) genes, 13 protein-coding genes, 2 ribosomal RNA (rRNA) genes and 1 non-coding control region. The overall base composition was e stimated to be A, 29.76%; T, 27.38%; C, 28.74% and G, 14.1% with GC content of 42.85%. This first published mitochondrial genome of Cynopterus allows further analysis of the species at a genomic scale for comparison with other mitochondrial genome of bats available. 21,22 D-Loop Repeatitive sequence at D-loop T A T GCG T A T GCG T A T GCG T A T GCG T A T GCG T A T GCG T A T GCG T A T GCG T A T GCG T A T GCG T A T GCG T A T GCG DNA sequencing of a long string of N(s) in D-loop / control region showed hexanucleotide repeats (TATGCG) 154 . Reference Bernt M, Donath A, Jühling F, Externbrink F, Florentz C, Fritzsch G, Pütz J, Middendorf, Stadler PF. 2013. MITOS: Improved de novo Metazoan Mitochondrial Genome Annotation. Molecular Phylogenetics and Evolution 69(2):313-319. Carver T, Harris SR, Berriman M, Parkhill J and McQuillan JA. 2012. Artemis: an integrated platform for visualization and analysis of high-throughput sequence-based experimental data. Bioinformatics (Oxford, England);28;4;464-9. Carver T, Thomson N, Bleasby A, Berriman M and Parkhill J. 2009. DNAPlotter: circular and linear iteractive genome visualization. Bioinformatics (Oxford England);25;1;119-20. Tamura K, Stecher G, Peterson D, Filipski A, and Kumar S. 2013. MEGA6: Molecular Evolutionary Genetics Analysis Version 6.0. Molecular Biology and Evolution 30: 2725-2729. Zerbino DR, Birney E. 2008. Velvet: algorithms for de novo short read assembly using de Bruijn graphs. Genome Research 18:821–829. doi:10.1101/gr.074492.107. Methodology Sampling 1 DNA extraction 2 NGS (Illumina Hiseq2000) 3 Assembly Velvet 1.2.10 4 Annotation MITOS revision 583 7 Interactive view DNAPlotter 10 Mapping reads ARTEMIS 14.0.0 9 Primer design flanking N reads Primer 3web 4.0 5 DNA sequencing 6 Phylogenomic analysis MEGA 6 8 Protein coding genes Non-protein coding genes tRNA genes Legend Nucleotide Composition 29.76% 28.74% 27.38% 14.10% 0.00% 5.00% 10.00% 15.00% 20.00% 25.00% 30.00% 35.00% A C T G List of tRNA genes and its products by order ( blue numbers ) No. Gene Product 1. trnF(gaa) tRNA-Phe 2. trnV(tac) tRNA-Val 3. trnL(taa) tRNA-Leu1 4. trnI(gat) tRNA-Ile 5. trnQ(ttg) tRNA-Gln 6. trnM(cat) tRNA-Met 7. trnW(tca) tRNA-Trp 8. trnA(tgc) tRNA-Ala 9. trnN(gtt) tRNA-Asp 10. trnC(gca) tRNA-Cys 11. trnY(gta) tRNA-Tyr 12. trnS(tga) tRNA-Ser1 13. trnD(gtc) tRNA-Asn 14. trnK(ttt) tRNA-Lys 15. trnG(tcc) tRNA-Gly 16. trnR(tcg) tRNA-Arg 17. trnH(gtg) tRNA-His 18. trnS(gct) tRNA-Ser2 19. trnL(tag) tRNA-Leu2 20. trnE(ttc) tRNA-Glu 21. trnT(tgt) tRNA-Thr 22. trnP(tgg) tRNA-Pro 3D image of tRNA CCA tail in orange Acceptor stem in purple D arm in red Anticodon arm in blue Anticodon in black Results Total length: 16848 bps No. protein coding genes: 13 No. Ribosomal genes: 2 No. tRNA genes: 22 D-Loop with TATGCG repeats tRNA genes not coded: Asx & Glx 4000 5000 6000 7000 8000 9000 10000 11000 12000 13000 14000 15000 16000 0 1000 2000 3000 *K2P NJ tree generated using bat’s mitochondrial genomes found in NCBI and C. sphinx in this study. 0.02 NC023122 Pteropus alecto KF726143 Pteropus alecto NC002612 Pteropus dasymallus` AB042770 Pteropus dasymallus NC002619 Pteropus scapulatus NC007393 Rousettus aegyptiacus AB205183 Rousettus aegyptiacus KF750629 UNVERIFIED Cynopterus sphinx SAP001 Cynopterus sphinx Malaysia NC005434 Rhinolophus pumilus NC018540 Hipposideros armiger NC022423 Desmodus rotundus NC022427 Sturnira tildae NC002009 Artibeus jamaicensis NC005436 Pipistrellus abramus NC015828 Myotis formosus NC005435 Sorex unguiculatus 92 100 100 100 100 100 100 100 100 100 100 100 100 89 Acknowledgements We would like to thank PERHILITAN for permit appoval to collect bats for research purpose, UMK and USM for various administrative and logistic support, and GENOM Malaysia for NGS facility acess. Special thanks to Puan Irni Suhayu Sapian for mentoring students in sample preparation. To Richard Teh Swee Aun and Siti Hajjar Mohd Daud, we are greatful for thier effort in the sample preparation procedure.