cole-trapnell-lab / cufflinks

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Getting warning while using cuffnorm - "Warning: Using default Gaussian distribution due to insufficient paired-end reads in open ranges. It is recommended that correct parameters (--frag-len-mean and --frag-len-std-dev) be provided." #112

Open sourav8888 opened 5 years ago

sourav8888 commented 5 years ago

**Hi, During cuffnorm for 18 samples run getting this warning (9 times and ):

Warning: Using default Gaussian distribution due to insufficient paired-end reads in open ranges. It is recommended that correct parameters (--frag-len-mean and --frag-len-std-dev) be provided.

I tried to investigate that myself and I got these:

Cufflink assembly stdout:**

**[14:43:30] Loading reference annotation. [14:43:30] Inspecting reads and determining fragment length distribution.

Processed 4057 loci. [] 100% Map Properties: Normalized Map Mass: 14847595.00 Raw Map Mass: 14847595.00 Fragment Length Distribution: Empirical (learned) Estimated Mean: 409.87 Estimated Std Dev: 150.14 [14:45:32] Assembling transcripts and estimating abundances. Processed 4057 loci. [] 100%**

**So here, fragment length mean and std-dev is calculated based on SAM alignment.

But while merging:**

[bam_header_read] EOF marker is absent. The input is probably truncated. [bam_header_read] invalid BAM binary header (this is not a BAM file). File assembled_unique/tmp/mergeSam_file8AmUL8 doesn't appear to be a valid BAM file, trying SAM... [04:06:36] Loading reference annotation. [04:06:36] Inspecting reads and determining fragment length distribution. Processed 1884 loci.

Map Properties: Normalized Map Mass: 252380.00 Raw Map Mass: 252380.00 Fragment Length Distribution: Truncated Gaussian (default) Default Mean: 200 Default Std Dev: 80 [04:06:37] Assembling transcripts and estimating abundances. Processed 1884 loci.

**So cuffmerge is applying default fragment length related parameter while merging, Thus cuffnorm is showing the warning I am getting.

I tried to define --frag-len-mean and --frag-len-std-dev to the any of these two steps, but didn't find any options. So, my question is whether it will cause any problem to the assembly and abundance calculation.**

Thanks!