We designed a pair of specific primers (Forward: 5′-GAG CCG ATT GTT GAA GCG GAA AAA G-3′; Reverse: 5′-TGG CCT TGA TAG CGT TGT TCA AAA T-3′) of Garfield_BM (Zhang et al. 2014 (link)) using its internal sequence to determine its distribution in some insects. PCR was performed with an initial denaturation step of 4 min at 95 °C followed by 30 cycles of 40 s at 95 °C, 40 s at 58 °C, and 2 m at 72 °C. Then, purified PCR products were cloned into PMD-19 cloning vector (TaKaRa). One or two random clones of each species were selected and sequenced.
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