PCR amplification and sequencing of rice-specific DNA barcodes
The PCR reaction mixture contained 1× Taq buffer with
Mg2+,
0.1 mM dNTPs, and 20 ng DNA. The PCR program included 40 cycles at 94°C
for 30 s, 55°C for 30 s, and 72°C for 2 min. PCR products were cleaned
using PEG8000 and sequenced in both directions on an ABI 3730xl DNA
Analyzer (Applied Biosystems). The sequences were assembled using
Sequencher 5.4 and edited if necessary to correct some nucleotide
calling mistakes.