# silva bacteria region postion

**URL:** <https://forum.mothur.org/t/silva-bacteria-region-postion/1875>\
**Category:** Commands in mothur\
**Created:** [May 29, 2014, 8:16pm UTC](https://forum.mothur.org/t/silva-bacteria-region-postion/1875 "2014-05-29T20:16:48Z")\
**Posts on this page:** 3\
**Page:** 1

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**Author:** ![jfgonzalezp](https://avatars.discourse-cdn.com/v4/letter/j/7cd45c/32.png) [@jfgonzalezp](https://forum.mothur.org/u/jfgonzalezp)\
**Post date:** [May 29, 2014, 8:16pm UTC](https://forum.mothur.org/t/silva-bacteria-region-postion/1875/1 "2014-05-29T20:16:48Z")

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Hi,

I’m not sure which region of the silva reference database my primers cover, they are 27F forward and 519R, I saw a previous entry in the forum which said to run pcr.seqs on silva.bacteria.fasta using an oligos file including both the forward and revers primers and setting keepdots=T, then running summary.seqs on the output file.

I followed this procedure but my output silva.bacteria.pcr.fasta file is empty, what can I be doing wrong?

I entered this command:

mothur \> pcr.seqs(fasta=silva.bacteria.fasta, oligos=pcrTest.oligos, processors=8)

Here is my oligos file:

forward AGRGTTTGATCMTGGCTCAG  
reverse GTNTTACNGCGGCKGCTG  
barcode CGTTTCTA N107  
barcode CTAACACA N115  
barcode CGTGATAA N118  
barcode CGTGGGAC N119  
barcode CGTCTGAA N12  
barcode CGTCGCAT N3  
barcode CGTGGTCA N37  
barcode CGTTCACG N54  
barcode CGTTTACT N56  
barcode CGTGAGAC N97  
barcode CGTAACCA P13  
barcode CGTATTTC P18  
barcode CGTAAGAA P23  
barcode CGTACCCA P36  
barcode CGTCAAGA P46  
barcode CGTAGATA P50  
barcode CGTAGGCT P61  
barcode CGTCACAG P75  
barcode CGTCCAGG P79  
barcode CGTATTCA P9

When I run summary files:

mothur \> summary.seqs(fasta=current)

I get this:

Using silva.bacteria.pcr.fasta as input file for the fasta parameter.  
[ERROR]: silva.bacteria.pcr.fasta is blank, aborting.  
Using silva.bacteria.pcr.fasta as input file for the fasta parameter.

Using 8 processors.  
[ERROR]: silva.bacteria.pcr.fasta is blank. Please correct.  
Error in reading your fastafile, at position -1. Blank name.

Thanks for your help!

Juan

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**Author:** ![pschloss](https://yyz2.discourse-cdn.com/flex036/user_avatar/forum.mothur.org/pschloss/32/4_2.png) [@pschloss](https://forum.mothur.org/u/pschloss)\
**Post date:** [May 30, 2014, 12:41pm UTC](https://forum.mothur.org/t/silva-bacteria-region-postion/1875/2 "2014-05-30T12:41:56Z")

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Hi Juan,

So when we built the silva.bacteria.fasta reference we removed the 27f and 1492r primer sequences since these aren’t really sequencable (unless you do genomes). So the start position will be 1044. If you remove that from your oligos file to run pcr.seqs and then run summary.seqs you’ll get 1044 and 13127 as your start and end positions.

Pat

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**Author:** ![jfgonzalezp](https://avatars.discourse-cdn.com/v4/letter/j/7cd45c/32.png) [@jfgonzalezp](https://forum.mothur.org/u/jfgonzalezp)\
**Post date:** [May 30, 2014, 2:47pm UTC](https://forum.mothur.org/t/silva-bacteria-region-postion/1875/3 "2014-05-30T14:47:30Z")

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Thanks Pat, that worked great!