TY - JOUR AU - SUMANTADINATA, KOMAR AU - SUBYAKTO, SLAMET AU - ALIMUDDIN, AU - RUSTIDJA, AU - JATI, M. SASMITO AU - FAIZAL, IRVAN AU - ALIAH, RATU SITI AU - TRIASTUTIK, GEMI PY - 2024/01/11 Y2 - 2024/03/29 TI - COMPARISON OF THREE DIFFERENT TECHNIQUES OF GENE TRANSFER IN HUMPBACK GROUPER ( ) CROMILEPTES ALTIVELIS JF - BIOTROPIA JA - BIOTROPIA VL - 18 IS - 1 SE - Research Paper DO - 10.11598/btb.2011.18.1.134 UR - https://journal.biotrop.org/index.php/biotropia/article/view/134 SP - AB - Humpback grouper is one of the most cultured fishes in Asia, including Indonesia. The<br />main problemfaced by humpback culture is its slow growth rate.One of themethods that will<br />be more effective and efficient to solve the problem is using transgenic technique. This study<br />was conducted to determine the effectiveness of transfection,microinjection and electroporation<br />techniques on gene transfer in humpback grouper. transfection was performed by<br />incubating sperm to the foreign DNA (pktBP-ktGH gene construct)-transfectant complex<br />solution, while was by injecting those complex solution into testis of mature males.<br />Microinjection was conducted in 2-4 cell stage embryos using 25 μg/ml of foreign DNA<br />solution, and duration of injection was 1, 2 and 3 seconds. Electroporation by 50 V, 30 ms of<br />pulse length, 5 of pulse number and 0.1 of pulse interval was performed to sperm using three<br />DNA concentration of 5, 10 and 20 μg/ml. The incorporation of foreign DNA in sperm and<br />embryos were analyzed using PCR method. Based on PCR analysis, an optimum DNA<br />concentration for electroporation was 10 μg/ml. Limited number of embryos could be<br />microinjected during 20-30 min to reach 2-4 cell stage. Microinjection for 1 second showed<br />higher survival rate of embryos, although none or very low number of larvae was hatched.<br />Transfast was an effective DNA delivery reagent for humpback grouper sperm. Foreign DNA<br />could be detected in sperm from two out of ten transfected fish at least 36 hours post<br />transfection (hpt). By transfection, foreign DNA was detected in sperm at 48 hpt 25 C<br />incubation temperature. Our study revealed that transfection, microinjection as well as<br />electroporation could be used as transgenesis methods in humpback grouper. By means of<br />simplicity and efficacy, however, electroporationwas an appropriate gene transfermethod.<br />o<br />In vitro<br />in vivo<br />in vivo<br />in vitro<br /> ER -