Warning!: all frames are zero

[valvanom]

I have installed snippy in mac OX intel 64 using brew. No issues checking dependencies, except that the bcftools version required was 1.7 and the one installed was 1.16. I installed version 1.7 from source into the usr/local and the snippy --check recognised all dependencies as OK. However after running snippy the program is aborted. The log file is below:

echo snippy 4.6.0

cd /Users/valvano/Genomes/snippy/AC011/AC011_ref

/usr/local/bin/snippy --outdir mysnps --force --ref AC011.gbk --R1 9278_AC011CR-1_1_trimmed.unmatched.fastq --R2 9278_AC011CR-1_2_trimmed.unmatched.fastq

samtools faidx reference/ref.fa

bwa index reference/ref.fa

[bwa_index] Pack FASTA... 0.05 sec [bwa_index] Construct BWT for the packed sequence... [bwa_index] 2.23 seconds elapse. [bwa_index] Update BWT... 0.03 sec [bwa_index] Pack forward-only FASTA... 0.03 sec [bwa_index] Construct SA from BWT and Occ... 0.57 sec [main] Version: 0.7.17-r1188 [main] CMD: bwa index reference/ref.fa [main] Real time: 2.924 sec; CPU: 2.908 sec

mkdir -p reference/genomes && cp -f reference/ref.fa reference/genomes/ref.fa

ln -sf reference/ref.fa .

ln -sf reference/ref.fa.fai .

mkdir -p reference/ref && gzip -c reference/ref.gff > reference/ref/genes.gff.gz

snpEff build -c reference/snpeff.config -dataDir . -gff3 ref

WARNING: All frames are zero! This seems rather odd, please check that 'frame' information in your 'genes' file is accurate.

bwa mem -Y -M -R '@RG\tID:mysnps\tSM:mysnps' -t 8 reference/ref.fa /Users/valvano/Genomes/snippy/AC011/AC011_ref/9278_AC011CR-1_1_trimmed.unmatched.fastq /Users/valvano/Genomes/snippy/AC011/AC011_ref/9278_AC011CR-1_2_trimmed.unmatched.fastq | samclip --max 10 --ref reference/ref.fa.fai | samtools sort -n -l 0 -T /private/var/folders/jz/j4h61y_135x8ymlqwvzmp50h0000gp/T --threads 3 -m 2000M | samtools fixmate -m --threads 3 - - | samtools sort -l 0 -T /private/var/folders/jz/j4h61y_135x8ymlqwvzmp50h0000gp/T --threads 3 -m 2000M | samtools markdup -T /private/var/folders/jz/j4h61y_135x8ymlqwvzmp50h0000gp/T --threads 3 -r -s - - > snps.bam

READ 0 WRITTEN 0 EXCLUDED 0 EXAMINED 0 PAIRED 0 SINGLE 0 DULPICATE PAIR 0 DUPLICATE SINGLE 0 DUPLICATE TOTAL 0

samtools index snps.bam

fasta_generate_regions.py reference/ref.fa.fai 457247 > reference/ref.txt

freebayes-parallel reference/ref.txt 8 -p 2 -P 0 -C 2 -F 0.05 --min-coverage 10 --min-repeat-entropy 1.0 -q 13 -m 60 --strict-vcf -f reference/ref.fa snps.bam > snps.raw.vcf

bcftools view --include 'FMT/GT="1/1" && QUAL>=100 && FMT/DP>=10 && (FMT/AO)/(FMT/DP)>=0' snps.raw.vcf | vt normalize -r reference/ref.fa - | bcftools annotate --remove '^INFO/TYPE,^INFO/DP,^INFO/RO,^INFO/AO,^INFO/AB,^FORMAT/GT,^FORMAT/DP,^FORMAT/RO,^FORMAT/AO,^FORMAT/QR,^FORMAT/QA,^FORMAT/GL' > snps.filt.vcf

Failed to open -: unknown file type

Any ideas? Thanks