Multiple Alignment and Primer Design for Groups of Barnacle Organisms and Amphibalanus amphitrite as Biofouling Markers

Abstract

. The impact caused by biota biofouling activities is the occurrence of corrosiveness and paralysis in installation buildings. To facilitate the installation and stability of biodiversity, researchers try to present environmental DNA (e-DNA) as environmental samples without killing and isolating an organism by utilizing environmental samples. This study aims to obtain a universal and specific primer of the Cytochrome Oxidase I (COX1) gene that can be used to identify biofouling barnacle groups designed in silico. The sequence used in this study was from Amphibalanus amphitrite mitochondria and Semibalanus balanoides with NCBI accession numbers NC_024525.1|:1-1,551 and NC_039849|:1-1,551. The sequence of COX1 sequences obtained was saved in FASTA format for further use in the primer design process using Geneious Prime. The determination of sustainable areas was determined using Benchling and Geneious Prime in the stages of multiple alignment. Sequence with the best specific primer criteria for COX1 A. amphitrite obtained with a length of 20-22 bases and an amplicon size of 910 bp. The sequence of forward and reverse primer bases were 5'-GAGCTGAACTTGGTCAACCG-3' and 5'GCTCAAAGAAGAGGAGGGCTAT-3'. Sequence with best universal primer criteria of the barnacle group for COX1 were obtained with a length of 24 bases each and an amplicon size of 1,270 bp. The sequence of forward and reverse primer bases were 5'-GACTTCTACCGTTAATGTTAGGAG-3' and 5'- CTATGGTAATGAGGAGGAGTAGTG-3'. The design results meet the requirements of a good primer so that the primer candidate design results can be used for the PCR process.