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Primer3 Input -version 0.4.0- -

| Tag | Default | Description | | --- | --- | --- | | PRIMER_TM_OPT | 60.0 | Optimal melting temp (°C) | | PRIMER_TM_MIN | 57.0 | Minimum acceptable Tm | | PRIMER_TM_MAX | 63.0 | Maximum acceptable Tm | | PRIMER_MAX_DIFF_TM | 100.0 | Max Tm difference between left and right primers |

-version 0.4.0- PRIMER_SEQUENCE_ID=COI_gene_fragment SEQUENCE=ATGGCTAGCTTAGCTAGCTAGCTAGCATCGATCGATCGATCGATCGATCGATCGATCGATCGATCGATCGATGC = primer3 input -version 0.4.0-

If you are attempting to run a command resembling primer3 input -version 0.4.0- , here are the most likely scenarios and how to fix them: | Tag | Default | Description | |

The settings are critical for preventing artifacts: primer3 input -version 0.4.0-

PRIMER_OPT_TM=60.0 PRIMER_MIN_TM=57.0 PRIMER_MAX_TM=63.0 PRIMER_MAX_DIFF_TM=3.0