Primer3 issue with the Sequence_ID
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12 days ago
aka ▴ 10

Hi everyone,

I am trying to sign primers for my sequences but I have an error that I do not understand the problem.

aka@aka:~/Desk/Primer\$ /home/aka/primer3/src/primer3_core < input.txt > result.txt


input.txt

PRIMER_TASK=generic
PRIMER_PICK_LEFT_PRIMER=1
PRIMER_PICK_INTERNAL_OLIGO=0
PRIMER_PICK_RIGHT_PRIMER=1
PRIMER_OPT_SIZE=20
PRIMER_MIN_SIZE=18
PRIMER_MAX_SIZE=23
PRIMER_MAX_NS_ACCEPTED=1
PRIMER_PRODUCT_SIZE_RANGE=100-150
P3_FILE_FLAG=1
PRIMER_EXPLAIN_FLAG=1
PRIMER_THERMODYNAMIC_PARAMETERS_PATH=/home/coralie/primer3/src/primer3_config/
=
SEQUENCE_ID=Qrob_P0702440.2
SEQUENCE_TEMPLATE=ATGCCTCCATCCTTCATCCACCGCCTACTCATCTCT....AATTGAGGAAGAGGGATTGTTCAATGAGTGGGAAGATTGCAAGACTTTGTTGA
=


It comes out: a results.txt file below, I feel like it's working but I have an error that shows up.

result.txt

    PRIMER_TASK=generic
PRIMER_PICK_LEFT_PRIMER=1
PRIMER_PICK_INTERNAL_OLIGO=0
PRIMER_PICK_RIGHT_PRIMER=1
PRIMER_OPT_SIZE=20
PRIMER_MIN_SIZE=18
PRIMER_MAX_SIZE=23
PRIMER_MAX_NS_ACCEPTED=1
PRIMER_PRODUCT_SIZE_RANGE=100-150
P3_FILE_FLAG=1
PRIMER_EXPLAIN_FLAG=1
PRIMER_THERMODYNAMIC_PARAMETERS_PATH=/home/coralie/primer3/src/primer3_config/
PRIMER_ERROR=Need PRIMER_SEQUENCE_ID if PRIMER_FILE_FLAG is not 0
=


When I change the SEQUENCE_ID to PRIMER_SEQUENCE_ID it doesn't work at all and shows me this:

PRIMER_TASK=generic
PRIMER_PICK_LEFT_PRIMER=1
PRIMER_PICK_INTERNAL_OLIGO=0
PRIMER_PICK_RIGHT_PRIMER=1
PRIMER_OPT_SIZE=20
PRIMER_MIN_SIZE=18
PRIMER_MAX_SIZE=23
PRIMER_MAX_NS_ACCEPTED=1
PRIMER_PRODUCT_SIZE_RANGE=100-150
P3_FILE_FLAG=1
PRIMER_EXPLAIN_FLAG=1
PRIMER_THERMODYNAMIC_PARAMETERS_PATH=/home/coralie/primer3/src/primer3_config/
PRIMER_ERROR=Need PRIMER_SEQUENCE_ID if PRIMER_FILE_FLAG is not 0
=
PRIMER_SEQUENCE_ID=Qrob_P0702440.2
SEQUENCE_TEMPLATE=ATGCCTCCATCCTTCATCCACCGCCTACTCATCTCT....AATTGAGGAAGAGGGATTGTTCAATGAGTGGGAAGATTGCAAGACTTTGTTGA
PRIMER_ERROR=Need PRIMER_SEQUENCE_ID if PRIMER_FILE_FLAG is not 0
=
PRIMER_SEQUENCE_ID=test2
SEQUENCE_TEMPLATE=GTAGTCAGTAGACNATGACNACTGACGATGCAGACNACACACACACACACAGCACACAGGTATTAGTGGGCCATTCGATCCCGACCCAAATCGATAGCTACGATGACG
PRIMER_ERROR=Need PRIMER_SEQUENCE_ID if PRIMER_FILE_FLAG is not 0
=
PRIMER_SEQUENCE_ID=test3
SEQUENCE_TEMPLATE=GTAGTCAGTAGACNATGACNACTGACGATGCAGACNACACACACACACACAGCACACAGGTATTAGTGGGCCATTCGATCCCGACCCAAATCGATAGCTACGATGACG
PRIMER_ERROR=Need PRIMER_SEQUENCE_ID if PRIMER_FILE_FLAG is not 0
=


Do you know where this error comes from? Are my primers good in the first case with the error?

Have a nice day

Aka

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