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Abstrak : Salah faktor penting yang diduga terlibat dalam penentuan jenis kelamin hewan TSD adalah Sox9. Protein ini berperan dalam cascade determinasi seks. Ketika kadar protein Sox9 mencapai nilai ambang tertentu maka gonad indiferen akan memasuki jalur diferensiasi testis. Penelitian ini bertujuan mengamati level ekspresi gen tersebut pada tahap awal perkembangan gonad Chelonia mydas yang termasuk dalam kelompok hewan TSD. Sebagian telur diinkubasi pada suhu 31±1°C untuk menghasilkan 100% individu betina (Suhu Feminisasi, SF), sedangkan telur lainnya diinkubasi pada suhu 26±1°C untuk menghasilkan 100% individu jantan (Suhu Maskulinisasi, SM). Pengamatan dilakukan pada level mRNA yang terekspresi sejak periode pra TSP hingga TSP akhir. Level ekspresi Sf1 kompleks adrenal-gonad-ginjal dari embrio tahap perkembangan 12, 14, 16 (pra TSP), 17, 19 dan 22 (TSP) yang diinkubasi pada suhu feminisasi (31±1°C) atau suhu maskulinisasi (26±1°C) dideteksi dengan menggunakan metode RT-PCR . Sebagai pembanding digunakan housekeeping gene â-actin . Hasil amplifikasi di elektroforesis pada Agarose-TAE kemudian di foto dengan kamera digital. Densitas larik DNA yang terbentuk pada masing-masing tahap perkembangan embrio dan perlakuan ditentukan dengan menggunakan software ScionImage. Rata-rata bacaan pada setiap larik di visualisasi dalam bentuk diagram garis. Hasil RT-PCR menunjukkan bahwa suhu inkubasi mempengaruhi level ekspresi Sox9 . Level ekspresi Sox9 tampak lebih tinggi pada embrio yang diinkubasi pada SM (26±1°C) dibandingkan dengan level ekspresi Sox9 embrio yang diinkubasi pada SF (31±1°C). Perbedaan level ekspresi tersebut terlihat sejak tahap pra TSP tengah hingga TSP tengah. Perbedaan terbesar terlihat menjelang embrio memasuki tahap TSP. Perbedaan level ekspresi Sox9 pada tahapan tersebut menunjukkan bahwa Sox9 memainkan peran yang penting dalam proses awal determinasi seks gonad Chelonia mydas . Kata kunci : Sox9, Temperature-dependent Sex Determination (TSD), Thermosensitive period (TSP), Chelonia mydas. Abstract : One of the important factor that is seems to be involved in sex determination of TSD animal is Sox9 . Sox9 has an important role in the molecular sex-determining cascade. When the level of Sox9 protein reach the treshold, it could drive the bipotential gonad to a testis differentiation . The research is conducted to examine Sox9 expression level in the early stage of gonadal development of Chelonia mydas , a species with TSD. Eggs were incubated in FPT (31±1°C) incubator to produce 100% of female embryo, and the remaining in MPT (26±1°C) produce 100% of male embryo. The examination level expression of mRNA was performed since preTSP till the end of TSP period. Quantification of Sf1 level expression from adrenal-gonad-kidney (AGK) complex of embryonic stage 12, 14, 16 (pre TSP), 17, 19 dan 22 (TSP), incubated in FPT (31±1°C) or MPT (26±1°C) were performed using RT-PCR method. Level expression of housekeeping gene â-actin is used as a control. Amplification product was electrophorized on TAE-Agarose gel and then documenting by using digital camera. The density of each DNA band were examined by using ScionImage Software. The average of 10 scores for each band was convert into a line diagram. RT-PCR result showed that the incubation temperature influenced mRNA expression level of Sox9. Expression level of Sox9 is higher in embryo which were incubated in MPT (26±1°C) than in FPT (31±1°C). Differential expression level of the gene appear from the middle of preTSP till the middle of TSP. The highest level of the difference appear soon before Thermosensitive Period. Differential expression of Sox9 during the stage showed that Sox9 has an important role during the early stage of sex determination gonad of Chelonia mydas . Key words: Sox9, Temperature-dependent Sex Determination (TSD), Thermosensitive period (TSP), Chelonia mydas

SUHU INKUBASI MEMPENGARUHI LEVEL EKSPRESI SOX9 PADA TAHAP AWAL PERKEMBANGAN GONAD EMBRIO Chelonia mydas (PENYU HIJAU)