Assembly Anomalies and Other Reasons a Genome Assembly may be Excluded from RefSeq

Introduction

A genome assembly may be excluded from the NCBI Reference Sequence (RefSeq) project for any of the reasons listed below. A few of these reasons for exclusion are likely to be of concern to all users, hence, we report these as "Assembly anomalies".

If the reason for exclusion from RefSeq would also impact your intended use of the genome assembly, you can apply filters to remove such assemblies from the search results (these filters appear in the left-hand sidebar under the heading "Exclude", filters for more reasons can be exposed using the "Customize …" menu). Filters can be applied for each individual reason, or the "excluded from RefSeq" property available under the "Advanced" search menu can be used to filter out all the genome assemblies that were excluded from RefSeq.

A filter to remove all assemblies flagged as anomalous from the search results in on by default. This filter must be cleared if you want to have assemblies flagged as anomalous included in the search results or if you want to filter out some types of anomaly but not others.

"Excluded from RefSeq" reasons can be reported for genome assemblies from any taxonomic group, however, many of the reasons for excluding an assembly from RefSeq are only revealed by running the NCBI Prokaryotic Genome Annotation Pipeline (PGAP).

Reasons an assembly is excluded from RefSeq

  • abnormal gene to sequence ratio - the ratio of the number of predicted genes to the length of the genome divided by 1000 is far outside the usual range for a Complete Genome assembly. The NCBI Prokaryotic Genome Annotation Pipeline typically expects to find an average of one gene for every 1,000 nucleotides in a genome assembly. The typical range is 0.8 to1.2; anything outside the range 0.5 to 1.5 is considered abnormal.
  • derived from metagenome - the genomic sequence was assembled from metagenomic sequencing rather than a pure culture leading to concerns about the accuracy of organism assignment and possible cross-contamination.
  • derived from environmental source - the source material for the assembly is from an environmental source rather than a pure culture leading to concern about the accuracy of organism assignment and possible cross-contamination.
  • derived from single cell - the source material for the assembly was amplified from a single cell leading to concern about the genome sequence accuracy.
  • derived from surveillance project - assembly generated from a pathogen surveillance project.
  • genome length too large - total non-gapped sequence length of the assembly is more than 1.5 times that of the average for the genomes in the Assembly resource from the same species, more than 15 Mbp, or is otherwise suspiciously long.
  • genome length too small - total non-gapped sequence length of the assembly is less than half that of the average for the genomes in the Assembly resource from the same species, less than 300 Kbp, or is otherwise suspiciously short.
  • high contig L50 - contig L50 is above 500.
  • low contig N50 - contig N50 is below 5000.
  • low gene count - the number of predicted genes is much lower than expected.
  • low quality sequence - long stretches of the sequence have a high proportion of ambiguous bases, are low complexity, or have some other indication that the sequence quality is low.
  • many frameshifted proteins - the CDSs predicted by the NCBI Prokaryotic Genome Annotation Pipeline have a suspiciously high number of frameshifts. For any clade containing at least 10 good quality assemblies the cutoff is more than three standard deviations from average or 5% of annotated CDSs, whichever is larger. For any clade containing less than 10 good quality assemblies the cutoff is more than 30% of total CDSs.
  • metagenome - a metagenome containing sequences from a mixture of organisms.
  • missing ribosomal protein genes - the NCBI Prokaryotic Genome Annotation Pipeline failed to find at least one copy of each essential ribosomal protein.
  • missing rRNA genes - the NCBI Prokaryotic Genome Annotation Pipeline failed to find at least one copy each of 5S, 16S, and 23S rRNA gene.
  • missing tRNA genes - the NCBI Prokaryotic Genome Annotation Pipeline failed to find tRNA genes with anticodons for 2 or more of the expected 20 amino-acids.
  • partial - the assembly has only partial genome representation.
  • untrustworthy as type - does not meet the criteria for a type strain assembly for use in ANI analysis.

Anomalous assemblies

  • chimeric - sequences from two different organisms are joined together.
  • contaminated - sequences from another organism, cloning vectors, linkers, adapters or primers are present in the assembly.
  • hybrid - sequences from a hybrid between different species, strains or isolates.
  • misassembled - alignment to related genome assemblies or other evidence indicates the assembly is likely to have errors.
  • mixed culture - sequences come from two or more organisms that were not cultured separately.
  • sequence duplications - assembly has one or more large duplications.
  • unverified source organism - the origin of the assembly is misidentified.

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Last updated: 2019-03-26T15:44:57Z