Designing Primers for Amplification of Coi, Cyt-B, Nd2 and Fib7 in Swiftlet (Aerodramus spp.)

Background: Edible bird’s nests (EBN) are highly nutritious and valued food for their numerous health benefits. Consequently, their high economic value and increasing global consumption demand for EBN have contributed to a rise in counterfeited food issue. Currently, the common method for identifying the origin of edible bird nests is the DNA barcoding PCR-based technique because of the limitations of morphological identification methods; however, the primers currently used for PCR face specific challenges in amplifying DNA from edible bird nests. Thus, newly designed primers are necessary. Methods: This study aims to redesign primers from the DNA barcode sequences of the mitochondrial genes cytochrome-c oxidase subunit I (COI), NADH dehydrogenase subunit II (ND2), cytochrome b (Cyt-b) and the nuclear gene â-fibrinogen intron 7 (Fib7) of Swiftlet belonging to the Aerodramus genus. Result: Four primer pairs AeroCOIf/AeroCOIr, AeroND2f/AeroND2r, AeroCytbf/AeroCytbr Cyt-b gene and AeroFib7f/AeroFib7r were designed successfully amplifying DNA from Aerodramus spp. The resulting amplicon sizes were 157 bp for COI, 150 bp for ND2, 130 bp for Cyt-b and 173 bp for Fib7 with the optimal annealing temperature of 60oC. For the result of the primers sensitivity investigation, the AeroCOIf/AeroCOIr, AeroND2f/AeroND2r, AeroCytbf/AeroCytbr and AeroFib7f/AeroFib7r primers was able to detect DNA concentration at 15´10-4 ng/µL, 15 ng/µL, 15´10-3 ng/µL and 15 ng/µL, respectively. These results demonstrate that these new primers could effectively amplify DNA from EBN belonging to Aerodramus spp.