The extent of common protein sequences in different assemblies and parameters from Japanese quail datasets against chicken proteome database.

<p>Panel A represents the intra-assembling parameterization of PAVE at 80% and 90% identity and Panel B represents the inter-assembling comparison among PAVE, MIRA and CAPRG. Panel C represents the intra-assembling parameterization of PAVE at 80% and 90% identity and Panel D represents the inter-assembling comparison among PAVE, MIRA and CAPRG.</p

CAPRG: Sequence Assembling Pipeline for Next Generation Sequencing of Non-Model Organisms

<div><p>Our goal is to introduce and describe the utility of a new pipeline “Contigs Assembly Pipeline using Reference Genome” (CAPRG), which has been developed to assemble “long sequence reads” for non-model organisms by leveraging a reference genome of a closely related phylogenetic relative. To facilitate this effort, we utilized two avian transcriptomic datasets generated using ROCHE/454 technology as test cases for CAPRG assembly. We compared the results of CAPRG assembly using a reference genome with the results of existing methods that utilize <em>de novo</em> strategies such as VELVET, PAVE, and MIRA by employing parameter space comparisons (intra-assembling comparison). CAPRG performed as well or better than the existing assembly methods based on various benchmarks for “gene-hunting.” Further, CAPRG completed the assemblies in a fraction of the time required by the existing assembly algorithms. Additional advantages of CAPRG included reduced contig inflation resulting in lower computational resources for annotation, and functional identification for contigs that may be categorized as “unknowns” by <em>de novo</em> methods. In addition to providing evaluation of CAPRG performance, we observed that the different assembly (inter-assembly) results could be integrated to enhance the putative gene coverage for any transcriptomics study.</p> </div

Assembly comparison for <i>Coturnix japonica</i> with various assembler programs and parameter spaces.

<p><i>E-value</i> cutoff for all database searches was <10E-05. The abbreviation “nr” represents non-redundant protein database from NCBI and “K” represents K-mer size. CAPRG* are assembly of reads that mapped to reference genome singly or failed to assemble by windowing against chromosome.</p

Runtime for each program.

<p>Assembly times represent execution on a computer with a duo 2.26 GHz Quad core Intel Xeon processor, 16 GB of RAM and 64 bit Snow Leopard v1.6 operating system.</p

Distribution of contigs per chromosome for Japanese quail and Northern bobwhite against the chicken reference genome.

<p>Distribution of contigs per chromosome for Japanese quail and Northern bobwhite against the chicken reference genome.</p

The effect of phylogenetic diversity on the assembling performance.

<p>Panel A Total number of reads mapped against chicken and zebra finch genome for <i>Coturnix japonica and Colinus virginianus</i>. Panel B Total number of unique hits against nr database for <i>Coturnix japonica and Colinus virginianus</i> mapped against chicken and zebra finch genome. <i>E-value</i> cutoff for all database searches was <10E-05. The abbreviation “nr” represents non-redundant protein database from NCBI.</p

The flow chart of CAPRG representing the mapping of reads to generate contiguous sequences (contigs).

<p>The flow chart of CAPRG representing the mapping of reads to generate contiguous sequences (contigs).</p

Assembly comparison for <i>Colinus virginianus</i> using various assembler programs and parameter spaces.

<p><i>E-value</i> cutoff for all database searches was <10E-05. The abbreviation “nr” represents non-redundant protein database from NCBI and “K” represents K-mer size. CAPRG* are assembly of reads that mapped to reference genome singly or failed to assemble by windowing against chromosome.</p