Desain dan Evaluasi in Silico Primer Spesifik Gen matK (maturase K) pada Genus Vaccinium Menggunakan Oligo Calc dan Primer-BLAST
DOI:
https://doi.org/10.59585/bajik.v4i2.1502Keywords:
Vaccinium, matK, Desain Primer, Oligo Calc, Primer-BLASTAbstract
Gen maturase K (matK) merupakan salah satu marka DNA barcoding standar pada tumbuhan tinggi yang memiliki tingkat evolusi ideal untuk analisis kloroplas, filogenetik, dan identifikasi spesies. Penelitian ini bertujuan untuk merancang dan mengevaluasi pasangan primer PCR secara in silico yang spesifik untuk amplifikasi gen matK pada genus Vaccinium. Metode yang digunakan meliputi penyelarasan sekuens (multiple sequence alignment) dari 10 sekuens gen matK genus Vaccinium yang diperoleh dari basis data NCBI, analisis rasio GC/AT, perancangan pasangan primer forward dan reverse, pemeriksaan struktur sekunder (seperti hairpin dan self-dimerization) dengan alat Oligo Calc, serta validasi spesifisitas amplikon menggunakan NCBI Primer-BLAST. Hasil analisis menunjukkan kandungan basa kelompok berada pada proporsi GC 40% dan AT 60%. Pasangan primer kandidat berhasil dirancang dan ditapis; kandidat primer yang membentuk struktur hairpin atau self-annealing dieliminasi. Evaluasi akhir menghasilkan dua set pasangan primer spesifik yang potensial untuk analisis PCR: Set Primer 1 (Forward: 5'-CATGTATGTGAATACGAATCTATCT-3', Reverse: 5'-GGCACTAGAGTATCGAACTTCT-3') menghasilkan amplikon spesifik berukuran 457 bp, serta Set Primer 2 (Forward: 5'-CAGCAACATGACTTCATATATCC-3', Reverse: 5'-TATGGATCCTTCTTGGTTGAGC-3')menghasilkan amplikon spesifik berukuran 222 bp pada plastida Vaccinium. Pasangan primer ini memenuhi syarat termodinamika dan menjadi kandidat kuat untuk aplikasi barcode DNA secara in vitro
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