<p>Tuna is the second largest fishery commodity in Indonesia after the shrimp. Since the<br />high demand and the limited stock of tuna resulted in fraudulent chance. Authentication<br />is required to meassure consumers regarding the accuracy of its labeling and food<br />safety. In this study, the authentication was based on protein and DNA barcoding using<br />cytochrome-b gene (cyt-b) of the mitochondrial DNA as the target of gene. Primer of<br />cyt b gene was designed based on the tuna species. This study aimed to identify the<br />authenticity of tuna fresh and its processed products through protein using SDS-PAGE </p><p>and DNA barcoding techniques. The phases of this research were protein electrophoresis<br />by SDS-PAGE, DNA extraction, PCR amplification, electrophoresis and sequencing.<br />Samples of fresh fish (Tu1, Tu2, Tu3, Tu4, and Tu5) and processed tuna (canned and<br />steak) were successfully extracted. Result showed that SDS-PAGE proved the damage of<br />proteins in the processed tuna, so this method was not appropriate if it is used to identify<br />the authenticity of tuna. PCR electrophoresis results showed that the samples of tuna,<br />tuna steak, sushi, meat ball, abon, and caned tuna were successfully amplified in the range<br />of 500-750 bp except Ka3, which was in line with the target of DNA (620 bp). Resulted<br />sequences of Tu2, Tu3, Tu4 and Tu5 were identified according the results of morphometric<br />namely T. albacares, while Tu1 was identified as T. obesus with homology level of 99%.<br />Processed tunas (steak and canned tuna) were identified as T. albacares, as stated on the<br />labels.<br />Keywords: Authentication, cytb, DNA barcoding, design primer, SDS-PAGE</p>