Table 2

PCR conditions for amplification of genomic DNA

PCR*Initial denaturationCycles; denaturationAnnealingExtensionIncubation
Region −8.8 94°C for 3 min 35 cycles; 98°C for 10 s 60°C for 15 s 68°C for 4 min 68°C for 7 min 
Region +5.8 94°C for 3 min 35 cycles; 98°C for 10 s 60°C for 15 s 68°C for 3 min 68°C for 7 min 
Region +11.5 94°C for 3 min 35 cycles; 98°C for 10 s 60°C for 15 s 68°C for 4 min 68°C for 7 min 
PCR1-PCR3 94°C for 3 min 35 cycles; 98°C for 10 s 60°C for 15 s 68°C for 16 min 68°C for 7 min 
PCR4 94°C for 3 min 35 cycles; 98°C for 10 s 66°C for 15 s 68°C for 8 min 68°C for 7 min 
PCR4 94°C for 3 min 35 cycles; 98°C for 10 s 66°C for 15 s 68°C for 1 min 68°C for 7 min 
PCR5 94°C for 3 min 35 cycles; 98°C for 10 s 66°C for 15 s 68°C for 8 min 68°C for 7 min 
PCR6 94°C for 3 min 35 cycles; 98°C for 10 s 66°C for 15 s 68°C for 6 min 68°C for 7 min 
PCR7 94°C for 3 min 35 cycles; 98°C for 10 s 66°C for 15 s 68°C for 10 min 68°C for 7 min 
PCR8 and PCR9 94°C for 3 min 30 cycles; 98°C for 1 min 55°C for 48 s 72°C for 3.4 min 72°C for 10 min 
PCR*Initial denaturationCycles; denaturationAnnealingExtensionIncubation
Region −8.8 94°C for 3 min 35 cycles; 98°C for 10 s 60°C for 15 s 68°C for 4 min 68°C for 7 min 
Region +5.8 94°C for 3 min 35 cycles; 98°C for 10 s 60°C for 15 s 68°C for 3 min 68°C for 7 min 
Region +11.5 94°C for 3 min 35 cycles; 98°C for 10 s 60°C for 15 s 68°C for 4 min 68°C for 7 min 
PCR1-PCR3 94°C for 3 min 35 cycles; 98°C for 10 s 60°C for 15 s 68°C for 16 min 68°C for 7 min 
PCR4 94°C for 3 min 35 cycles; 98°C for 10 s 66°C for 15 s 68°C for 8 min 68°C for 7 min 
PCR4 94°C for 3 min 35 cycles; 98°C for 10 s 66°C for 15 s 68°C for 1 min 68°C for 7 min 
PCR5 94°C for 3 min 35 cycles; 98°C for 10 s 66°C for 15 s 68°C for 8 min 68°C for 7 min 
PCR6 94°C for 3 min 35 cycles; 98°C for 10 s 66°C for 15 s 68°C for 6 min 68°C for 7 min 
PCR7 94°C for 3 min 35 cycles; 98°C for 10 s 66°C for 15 s 68°C for 10 min 68°C for 7 min 
PCR8 and PCR9 94°C for 3 min 30 cycles; 98°C for 1 min 55°C for 48 s 72°C for 3.4 min 72°C for 10 min 
*

Each reaction was performed by the 3-step PCR method. The PCR was started from the initial denaturation, followed by cycling reactions and incubation.

Alternative condition of PCR4 was used for direct sequencing of the shorter product derived from the Bm allele.

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