Media Peternakan, August 2012, pp. 102-110 Online version: EISSN 2087-4634 4ñ&&¥¨´∑¨ªï±∂ºπµ®≥ï∞∑©ï®™ï& 1 & Accredited by DGHE No: 66b/DIKTI/Kep/2011 #.(ñ WVï[Y_^ ¥¨´∑¨ªïXVWXïY[ïXïWVX The Characteristics of Cytochrome C Oxidase Gene Subunit I in Wild Silkmoth Cricula trifenestrata Helfer and Its Evaluation for Species Marker Surianaa *, D. D. Solihinb #, R. R. Noorc #, & A. M. Thoharid a1#¨∑®πª¥¨µª11!∞∂≥∂Æ¿ð1%®™º≥ª¿112™∞¨µ™¨ð1'®≥º∂≥¨∂14µ∞Ω¨π∫∞ª¿ )≥µï1'$ 1,∂≤∂´∂¥∑∞ª1*®¥∑º∫1'∞±®º1!º¥∞13Ø∞´®π¥®1 µ´º∂µ∂غ1*¨µ´®π∞ð1(µ´∂µ¨∫∞® b#¨∑®πª¥¨µª11!∞∂≥∂Æ¿ð1%®™º≥ª¿112™∞¨µ™¨ð1!∂Æ∂π1 Æπ∞™º≥ªºπ®≥14µ∞Ω¨π∫∞ª¿ c#¨∑®πª¥¨µª11 µ∞¥®≥1/π∂´º™ª∞∂µ1®µ´13¨™Øµ∂≥∂Æ¿ð1%®™º≥ª¿11 µ∞¥®≥12™∞¨µ™¨ð1!∂Æ∂π1 Æπ∞™º≥ªºπ®≥14µ∞Ω¨π∫∞ª¿ #)≥µï1 Æ®ª∞∫1*®¥∑º∫1(/!1#®π¥®Æ®ð1!∂Æ∂π1W\\^Vð1(µ´∂µ¨∫∞® d#¨∑®πª¥¨µª11"∂µ∫¨πΩ®ª∞∂µð1%®™º≥ª¿11%∂π¨∫ªπ¿ð1!∂Æ∂π1 Æπ∞™º≥ªºπ®≥14µ∞Ω¨π∫∞ª¿ )≥µï1+∞µÆ≤®π1 ≤®´¨¥∞≤1#®π¥®Æ®ð1!∂Æ∂π1W\\^Vð1(µ´∂µ¨∫∞® (Received 21-02-2012; accepted 23-04-2012) ABSTRAK Penelitian bertujuan untuk mengkarakterisasi dan mendeteksi situs diagnostik dari parsial gen sitokrom C oksidase sub unit I (COI) ulat sutera liar Cricula trifenestrata, dan mengevaluasi gen tersebut sebagai penanda spesies. Sebanyak 15 larva C. tifenestrata dikoleksi dari Kabupaten Bogor, Purwakarta, dan Bantul. DNA genom diekstrak dari kelenjar sutera larva, kemudian diperbanyak dengan metode PCR dan disekuensi. Hasil sekuensi dikarakterisasi runutan nukleotida dan diprediksi kandungan asam aminonya. Hasilnya menunjukkan bahwa 595 runutan nukleotida pada ujung 5’ gen COI C. tifenestrata bersifat kekal pada level spesies, tetapi beragam pada level family. Nukleotida didominasi oleh basa timin dan adenin (± 70%). Terdapat 25 situs diagnostik bagi C. tifenestrata dan 4 situk diagnostik pada level genus. Seratus delapan puluh sembilan (189) asam amino yang disejajarkan, menunjukkan satu persen variasi antar spesies. Asam amino ke-107 (valin) dan asam amino ke-138 (treonin) merupakan asam amino diagnostik bagi C. tifenestrata. Berdasarkan πºµºª®µ µº≤≥¨∂ª∞´® ´®µ ®∫®¥ ®¥∞µ∂µ¿®ð ≥∂ƨµ∞ ≤¨≤¨π®©®ª®µ ¥¨µºµ±º≤≤®µ ® C. tifenestrata terdapat pada nodus yang sama dengan spesies Antheraea, sehingga family Saturniidae bersifat ≥¨ª∞≤ï Kata kunci: ulat sutera liar, C. trifenestrata, sitokrom oksidase sub unit I, nukleotida diagnostik, asam amino diagnostik ABSTRACT The study was conducted to assess the characteristics of partial gene of cytochrome C oxidase subunit I (COI) of wild silkmoth Cricula trifenestrata, and to detect the diagnostic sites from these ƨµ¨ ¨Ω®≥º®ª∞∂µ ®∫ ∫∑¨™∞¨∫ ¥®π≤¨πï ª∂ª®≥ ≠ª¨¨µ ≥®πΩ®¨ C. tifenestrata were collected from Bogor, Purwakarta, and Bantul Regencies. Genomic DNA was extracted from silk gland of individual larvae, ªØ¨µ ®¥∑≥∞¨´ ©¿ /"1 ¥¨ªØ∂´ ®µ´ ∫¨∏º¨µ™¨´ï #- ∫¨∏º¨µ™∞µÆ ®∫ ´∂µ¨ ™Ø®π®™ª¨π∞¡¨ their nucleotide and amino acid contents. The results showed that 595 nucleotides at the 5 ‘end of COI gene of C. tifenestrata was conserved at the species level, but varies at the family level. Nucleotide dominated by thymine and adenine bases (± 70%). There were 25 diagnostic sites for C. tifenestrata, and four diagnostic sites for genus level. One hundred eigthty nine (189) amino acids were alignment, and only one percent of the genes was varied among species. The 107th amino acid (valine) and 138th (threonine) were diagnostics amino acid for C. tifenestrata. Based on nucleotides and amino acids ∫¨∏º¨µ™¨∫ðªØ¨ ¢≥∂ƨµ¿ ¨´ ªØ®ª C. tifenestrata lied on the same nodes with Antheraea, so the Saturniidae family is monophyletic. Key words: wild silk moth, C. trifenestrata, cytohrome C oxidase subunit I, nucleotide diagnostic, amino acid diagnostic * Corresponding author: $,¥®∞≥ñ[email protected] 102 August 2012 241( - 1ET AL. Media Peternakan INTRODUCTION 3Ø∞∫1∫ªº´¿1®∞¥¨´1ñ1ûWü1™Ø®π®™ª¨π∞¡¨ð1ûXü1´¨ª¨™ª1ªØ¨1 ∫∞ª¨11µº™≥¨∂ª∞´¨1®µ´1®¥∞µ∂1®™∞´1´∞®Æµ∂∫ª∞™∫1".(1ƨµ¨1 3ب1Æ∂≥´¨µ1∫∞≥≤¥∂ªØð11∂µ¨1≥∂™®≥1µ®¥¨11Cricula 1 C. trifenestratað1 ®µ´1 ûYü1 ¨ø®¥∞µ¨1 ªØ¨1 π¨≥®ª∞∂µ∫Ø∞∑1 1 trifenestrata '¨≥≠¨πò1 ®µ´1 ªØ¨1 ¥¨¥©¨π1 1 ªØ¨1 2®ªºπµ∞∞´®¨1 ≤∞µ∫Ø∞∑1 ®¥∂µÆ1 ªØ¨1 ∫∞≥≤,∑π∂´º™∞µÆ1 ∞µ∫¨™ª∫1 ©®∫¨´1 1 ≠®¥∞≥¿1 û∫º∑¨π≠®¥∞≥¿1 !∂¥©¿™∂∞´¨®üð1 1 1 ∞µ™≥º´¨1 µº™≥¨∂ª∞´¨1 ®µ´1 ®¥∞µ∂1 ®™∞´1 ".(1 ∫¨∏º¨µ™¨∫ï1 3ب1 π¨∫º≥ª∫1 ªØ¨1 Æ∞®µª1 ∫∞≥≤¥∂ªØ∫ð1 ¢®≥1 ð1 ®µ´1 ¨¥∑¨π∂π1 ï1 ¨π¨1 ´¨∫™π∞©¨´1 ©®∫¨´1 1 ƨµ¨ª∞™∫1 ©®™≤Æπ∂ºµ´∫ð1 1 1 3Ø∞∫1 ≠®¥∞≥¿1 1 ªØ¨1 ≥®πƨ∫ª1 ®µ´1 1 ∫∑¨™ª®™º≥®π1 1 ªØ¨1 1©¨1ªØ¨11¥∂≥¨™º≥®π1´®ª®©®∫¨11C. trifenestrata +¨∑∞´∂∑ª¨π®ð1 1 ®µ1 ¨∫ª∞¥®ª¨´1 µº¥©¨π1 1 WðYVV.Wð[VV1 1™®µ1©¨1º∫¨´1®∫1∫∑¨™∞¨∫1¥®π≤¨πï1 ∫∑¨™∞¨∫1 û3º∫≤¨∫1 et al.ð1 W__\üï1 3ب1 2®ªºπµ∞∞´®¨1 ≠®¥- ∞≥¿ð1 1 ¨ø®¥∑≥¨1 ´∂¥¨∫ª∞™®ª¨´1 ∫∞≥≤¥∂ªØ1 ûBombyx moriüð1 MATERIALS AND METHODS Ø®∫1 ®1 Æ𨮪1 ®4𮙪∞∂µ1 ®¥∂µÆ1 ªØ¨1 ∫∞≥≤¥∂ªØ1 ≠®¥∞≥¿1 û!∂¥©¿™∞´®¨üï1 (µ1 ®´´∞ª∞∂µð1 ∫∂¥¨1 ∫∑¨™∞¨∫1 1 ªØ¨1 ≠®¥∞≥¿1 1 ª∂ª®≥1 1 ≠ª¨¨µ1 ≥®πΩ®¨1 1 ªØ¨1 1 ∫∞≥≤¥∂ªØ1 C. ®π¨1 1 ∫∞≥≤¥∂ªØ∫1 1 ™®µ1 ®≥∫∂1 ∑π∂´º™¨1 ∫∞≥≤ï1 C. tri- trifenestrata1 ¨π¨1 ™∂≥≥¨™ª¨´1 1 !∂Æ∂π1 ®µ´1 ®≤®πª®1 fenestrata1 ®∫1™∂µ∫∑∞™º∂º∫1©¿1Ø®Ω∞µÆ1∑¨π≠∂𮪨´1™∂™∂∂µð1 1¨Æ¨µ™∞¨∫116¨∫ª1)®Ω®ð1®µ´1!®µªº≥11¨Æ¨µ™¿11"¨µªπ®≥1 1Æ∂≥´¨µ1¢ 1™∂≥∂πï1,®≥¨1 1∫∑®µ11[\,]^1ð1 Java, Indonesia. 1≠¨¥®≥¨1 1∫∑®µ11][,^W11û*®≤®ª∞1í1"غª∞®ð1 XVV_üï13Ø∞∫1∫∑¨™∞¨∫11´∞∫ªπ∞©ºª¨´11ªØπ∂ºÆØ∂ºª1%∞≥∞∑∞µ¨ð1 Genomic DNA Isolation (µ´∞®1®µ´1(µ´∂µ¨∫∞®ð1®µ´1ªØ¨π¨1®π¨1¥®µ¿1¨µ´¨¥∞™1∫∑¨- ™∞¨∫11(µ´∂µ¨∫∞®1(∫≥®µ´1∫º™Ø1®∫112º¥®ª¨π®ð1)®Ω®ð1®µ´1 &¨µ∂¥∞™1 #- 1 ®∫1 ∑π¨∑®π¨´1 1 ®1 ∑®∞π1 1 ∫∞≥≤1 2º≥®æ¨∫∞1 û-®∫∫∞Æ1 et al.,1 XVV\üï1 2ªº´¿1 ©®∫¨´1 1 ©∞∂≥∂Æ¿ð1 Æ≥®µ´∫1 1 µ®≥1 ∞µ∫ª®π1 ≥®πΩ®¨1 1 C. trifenestrata using a ¨™∂≥∂Æ¿1 ®µ´1 ƨµ¨ª∞™∫1 ®∫∑¨™ª∫1 1 1 ∞µ∫¨™ª1 1 ªØ¨π¨≠∂π¨1 ∫ª®µ´®π´1ª¨™Øµ∞∏º¨1û2®¥©π∂∂≤1et alïð1W_^_ü1ªØ®ª1Ø®∫1©¨¨µ1 necessary. As C. trifenestrata is a closely related to spe- ®´±º∫ª¨´ï ™∞¨∫11Antheraea and B. morið1ªØ¨1¥∞ª∂™Ø∂µ´π∞®≥1ƨµ∂¥¨1 ∞µ≠∂𥮪∞∂µ1 1 ªØ¨1 ∫∑¨™∞¨∫1 ¥®¿1 ∑π∂Ω∞´¨1 ≠ºµ´®¥¨µª®≥1 Electrophoresis ∞µ≠∂𥮪∞∂µ11≠ºªºπ¨1¢≥∂ƨµ¨ª∞™1®µ®≥¿∫¨∫1®µ´1¨Ω∂≥º- tionary biology. &¨µ∂¥∞™1#- 1 ®∫1¥∞Æ𮪨´11WïX–1®Æ®π∂∫¨1ƨ≥11 1¨™¨µª1¨∫ª∞Æ®ª∞∂µ∫1Ø®Ω¨1∫ºÆƨ∫ª¨´1¥∞ª∂™Ø∂µ´π∞®≥1 Wø3!$1∫∂≥ºª∞∂µ1º∫∞µÆ1ª∂∂≥∫11∫º©¥®π∞µ¨1¨≥¨™ªπ∂∑Ø∂π¨∫∞∫1 ƨµ∂¥¨1û¥ª#- ü1®∫1®1ª∂∂≥11∫ªº´¿∞µÆ1ªØ¨1ª®ø∂µ∂¥¿ð1 û'∂¨≠¨πð142 üï1$ªØ∞´∞º¥1©π∂¥∞´¨1ûVï[1ºÆ&¥≥ü1 ®∫1º∫¨´1 ®µ´1 ¨Ω∂≥ºª∞∂µ1 1 ®µ∞¥®≥1 ∑∂∑º≥®ª∞∂µ∫ï1 3ب1 ™¿ª∂™Øπ∂¥¨1 1∫ª®∞µ∞µÆ11ªØ¨1ƨ≥ð1®µ´1Ω∞∫º®≥∞¡¨´1ºµ´¨π1451ªπ®µ∫∞- ∂ø∞´®∫¨1 ∫º©ºµ∞ª1 (1 û".(ü1 ƨµ¨1 1 ®1 ∑®πª1 1 ¥ª#- ï1 ≥º¥∞µ®ª∂π1û•½1YVV1üï11 3Ø∞∫1 ƨµ¨1 ®∫1 ™Ø∂∫¨µ1 ©¨™®º∫¨1 1 1 ™¨µªπ®≥1 π∂≥¨1 1 ¥¨ª®©∂≥∞∫¥1 ®µ´1 1 ∑π¨∫¨µ™¨1 1 1 ®≥¥∂∫ª1 ®≥≥1 eukaryotes. ".( &¨µ¨ ¥∑≥∞™®ª∞∂µ ®µ´ 2¨∏º¨µ™∞µÆ ´´∞ª∞∂µ®≥≥¿ð1 ªØ¨1 £1 ®µ´1 ∫ªπº™ªºπ¨1 1 ".(1 ƨµ¨1 Ø®∫1 ©¨¨µ1 ¨≥≥1 ™∂µ∫¨πΩ¨´1 1 ªØ¨1 ®µ∞¥®≥1 Æπ∂º∑∫1 ®µ®≥¿∫¨´1 1 1 1π¨Æ∞∂µ11".(1ƨµ¨1´¨π∞Ω¨´11C. trifenestrata ≠®πð1®1≠¨®ªºπ¨1 1¥®≤¨∫11¨∫∑¨™∞®≥≥¿1∫º∞ª®©≥¨11¨Ω∂- ®∫1 ®¥∑≥∞¨´1 ©¿1 º∫∞µÆ1 ®1 3بπ¥®≥1 "¿™≥¨π1 $∑∑¨µ´∂π≠1 ≥ºª∞∂µ®π¿1∫ªº´∞¨∫1û+ºµª1et alïð1W__\üï13ب1".(ð1 ¨Ω¨πð11 3¿∑¨1 [YYX1 ¥®™Ø∞µ¨ï1 3ب1 ∑π∞¥¨π1 º∫¨´1 ¨π¨1 ®π´1 [, ∂µ¨11ªØ¨11™∂µ∫¨πΩ¨´1¥∞ª∂™Ø∂µ´π∞®≥1∑π∂ª¨∞µ,™∂´∞µÆ1 3& 3" , 333 3 3 ",Y1®µ´1π¨Ω¨π∫¨1[, 3 31 ƨµ¨∫1 1 ®µ∞¥®≥∫1 û,º¨≥≥¨πð1 XVV\üð1 ®µ´1 ªØº∫1 ´∞∫∑≥®¿∫1 ®1 3 ,3 3 ",Y1û,®Ø¨µ´π®µ1et al., XVV\üï11 ©¨4¨π1¢≥∂ƨµ¨ª∞™1∫∞Ƶ®≥1û6∞≥∫∂µð1XVWVüï1".(1®∫1®1©®π- 13ب1∑π∂Æπ®¥11/"11®¥∑≥∞™®ª∞∂µ1 ®∫1®∫1 1W1 ™∂´¨1 ¥®π≤¨π1 Ø®∫1 ©¨¨µ1 º∫¨´1 ¨π¿1 ∫º™™¨∫∫≠º≥≥¿1 1 ¥®µ¿1 1®ª1_Z17"ð1 ¨´1©¿1Y[1™¿™≥¨∫11YV1∫¨™1®ª1_Z17"ð1Z[1 ®µ∞¥®≥1 taxað1 1 ¨ø®¥∑≥¨∫ñ1 ¨π¨µª∞®ª¨´1 ®¥∂µÆ1 ≠®¥∞≥¿1 ∫¨™1®ª1ZV7"ð1®µ´1W11®ª1]X17"ð1 1®1∫º©∫¨∏º¨µª1[11 1+¨∑∞´∂∑ª¨π®1û6∞≥∫∂µð1XVWVüð1©¨¨ª≥¨∫1û%ºµ≤1et al.ð1W__[üð1 1 µ®≥1 ¨øª¨µ∫∞∂µ1 ®ª1 ]X1 7"ï1 1¨®Æ¨µª1 ™∂¥∑∂µ¨µª∫1 ¨π¨1 ∫∂¥¨1 ∞µ∫¨™ª1 ∑¨∫ª∫1 û3∂´®1 í1 ,ºπ®∞ð1 XVV\ü1 Mermerodes X1 •≥1 #- 1 ª¨¥∑≥®ª¨ð1 Wï[1 •≥1 1 ¨®™Ø1 WV1 ∑∞™∂¥∂≥1 ∑π∞¥¨πð1 hamona11û'º≥π™1et al.ð1XVV]üð1¥∂∫∏º∞ª∂1û"¿æ∞µ∫≤®1et WXï[1•≥11©º¨π1π¨®´¿11¡ð1Vï[1º≥1,Æ"≥X1[V¥,ð1®µ´1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 al.ð XVV\üð ®µ´ Thrips tabaci û*®π∞¥∞ et al., XVWVüï added double destiled H2.