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High-coverage sequencing and annotated assemblies of the budgerigar genome.

Ganapathy G, Howard JT, Ward JM, Li J, Li B, Li Y, Xiong Y, Zhang Y, Zhou S, Schwartz DC, Schatz M, Aboukhalil R, Fedrigo O, Bukovnik L, Wang T, Wray G, Rasolonjatovo I, Winer R, Knight JR, Koren S, Warren WC, Zhang G, Phillippy AM, Jarvis ED - Gigascience (2014)

Bottom Line: The sequence reads for this project were in part generated and used for both the Assemblathon 2 competition and the first de novo assembly of a giga-scale vertebrate genome utilizing PacBio single-molecule sequencing.Across several quality metrics, these budgerigar assemblies are comparable to or better than the chicken and zebra finch genome assemblies built from traditional Sanger sequencing reads, and are sufficient to analyze regions that are difficult to sequence and assemble, including those not yet assembled in prior bird genomes, and promoter regions of genes differentially regulated in vocal learning brain regions.This work provides valuable data and material for genome technology development and for investigating the genomics of complex behavioral traits.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Neurobiology, Duke University Medical Center, Durham, NC 27710, USA.

ABSTRACT

Background: Parrots belong to a group of behaviorally advanced vertebrates and have an advanced ability of vocal learning relative to other vocal-learning birds. They can imitate human speech, synchronize their body movements to a rhythmic beat, and understand complex concepts of referential meaning to sounds. However, little is known about the genetics of these traits. Elucidating the genetic bases would require whole genome sequencing and a robust assembly of a parrot genome.

Findings: We present a genomic resource for the budgerigar, an Australian Parakeet (Melopsittacus undulatus) -- the most widely studied parrot species in neuroscience and behavior. We present genomic sequence data that includes over 300× raw read coverage from multiple sequencing technologies and chromosome optical maps from a single male animal. The reads and optical maps were used to create three hybrid assemblies representing some of the largest genomic scaffolds to date for a bird; two of which were annotated based on similarities to reference sets of non-redundant human, zebra finch and chicken proteins, and budgerigar transcriptome sequence assemblies. The sequence reads for this project were in part generated and used for both the Assemblathon 2 competition and the first de novo assembly of a giga-scale vertebrate genome utilizing PacBio single-molecule sequencing.

Conclusions: Across several quality metrics, these budgerigar assemblies are comparable to or better than the chicken and zebra finch genome assemblies built from traditional Sanger sequencing reads, and are sufficient to analyze regions that are difficult to sequence and assemble, including those not yet assembled in prior bird genomes, and promoter regions of genes differentially regulated in vocal learning brain regions. This work provides valuable data and material for genome technology development and for investigating the genomics of complex behavioral traits.

No MeSH data available.


Related in: MedlinePlus

The distribution of read lengths in 454, Illumina and PacBio budgerigar sequences. The reads are binned into 5 bp buckets based on their lengths, and the fraction of reads (normalized by the size of the largest bucket) falling into each bucket is shown. Thus, curves shifted towards the right indicate longer read lengths. The reads labeled “20 Kbp”, “8 Kbp” and “3 Kbp”, “FLX Titanium” and “FLX Titanium XL+” are 454 reads. The reads labeled “PacBio pre-release C2” are uncorrected PacBio reads. The Illumina read lengths appear as colored square boxes, since these read lengths are uniform. The “Illumina Duke” reads are of length 76, The “Illumina UK” reads are of length 101, and the “Illlumina BGI” reads are of lengths 90 or 150. The longest reads come from PacBio sequencing, followed by 454 FLX + (i.e., FLX Titanium XL+) sequencing.
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Figure 1: The distribution of read lengths in 454, Illumina and PacBio budgerigar sequences. The reads are binned into 5 bp buckets based on their lengths, and the fraction of reads (normalized by the size of the largest bucket) falling into each bucket is shown. Thus, curves shifted towards the right indicate longer read lengths. The reads labeled “20 Kbp”, “8 Kbp” and “3 Kbp”, “FLX Titanium” and “FLX Titanium XL+” are 454 reads. The reads labeled “PacBio pre-release C2” are uncorrected PacBio reads. The Illumina read lengths appear as colored square boxes, since these read lengths are uniform. The “Illumina Duke” reads are of length 76, The “Illumina UK” reads are of length 101, and the “Illlumina BGI” reads are of lengths 90 or 150. The longest reads come from PacBio sequencing, followed by 454 FLX + (i.e., FLX Titanium XL+) sequencing.

Mentions: DNA samples were obtained from a blood sample taken from a single male Melopsittacus undulatus, who we aptly named Mr. B. For Illumina sequencing, reads were generated at Duke University (16×), Illumina UK (54×), and BGI (219×) using Illumina’s TruSeq [1] version2 or version3 chemistries (Table 1 and GigaDB [2]). The version3 chemistry reads through GC-rich regions, which are often found in promoters, more evenly than does version2 [3]. The insert sizes for the BGI libraries ranged from 220 bp to 40 Kbp, and the insert sizes for the Duke libraries ranged from 400–600 bp, in order to assist assemblies. Fragment sizes for the mate pair libraries, based on genome mapping, and the per base sequence quality distribution for the libraries are shown in GigaDB [2]. The Duke University Illumina libraries were sequenced at two different cluster densities: 8× coverage reads at the normal 420 k clusters/mm density and 8× coverage at a lower 350 k clusters/mm. The lower cluster density was used to increase the number of GC-rich regions sequenced. For PacBio sequencing, 6.76 Gbp (~5.5× coverage) of PacBio RS reads [4] were generated at Pacific Biosciences from two insert size libraries (7.5 K bp at 1.93× and 13 Kbp at 3.56×; PacBio reads error-corrected with Illumina can be downloaded from the supplementary webpage associated with [5]). With all reads combined, the total coverage exceeds 300× (assuming a haploid genome size of 1.23 Gbp) (Table 1), perhaps making Mr. B one of the most sequenced individual vertebrate animals as of to date. The read length distributions of these different types of reads are shown in Figure 1.


High-coverage sequencing and annotated assemblies of the budgerigar genome.

Ganapathy G, Howard JT, Ward JM, Li J, Li B, Li Y, Xiong Y, Zhang Y, Zhou S, Schwartz DC, Schatz M, Aboukhalil R, Fedrigo O, Bukovnik L, Wang T, Wray G, Rasolonjatovo I, Winer R, Knight JR, Koren S, Warren WC, Zhang G, Phillippy AM, Jarvis ED - Gigascience (2014)

The distribution of read lengths in 454, Illumina and PacBio budgerigar sequences. The reads are binned into 5 bp buckets based on their lengths, and the fraction of reads (normalized by the size of the largest bucket) falling into each bucket is shown. Thus, curves shifted towards the right indicate longer read lengths. The reads labeled “20 Kbp”, “8 Kbp” and “3 Kbp”, “FLX Titanium” and “FLX Titanium XL+” are 454 reads. The reads labeled “PacBio pre-release C2” are uncorrected PacBio reads. The Illumina read lengths appear as colored square boxes, since these read lengths are uniform. The “Illumina Duke” reads are of length 76, The “Illumina UK” reads are of length 101, and the “Illlumina BGI” reads are of lengths 90 or 150. The longest reads come from PacBio sequencing, followed by 454 FLX + (i.e., FLX Titanium XL+) sequencing.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4109783&req=5

Figure 1: The distribution of read lengths in 454, Illumina and PacBio budgerigar sequences. The reads are binned into 5 bp buckets based on their lengths, and the fraction of reads (normalized by the size of the largest bucket) falling into each bucket is shown. Thus, curves shifted towards the right indicate longer read lengths. The reads labeled “20 Kbp”, “8 Kbp” and “3 Kbp”, “FLX Titanium” and “FLX Titanium XL+” are 454 reads. The reads labeled “PacBio pre-release C2” are uncorrected PacBio reads. The Illumina read lengths appear as colored square boxes, since these read lengths are uniform. The “Illumina Duke” reads are of length 76, The “Illumina UK” reads are of length 101, and the “Illlumina BGI” reads are of lengths 90 or 150. The longest reads come from PacBio sequencing, followed by 454 FLX + (i.e., FLX Titanium XL+) sequencing.
Mentions: DNA samples were obtained from a blood sample taken from a single male Melopsittacus undulatus, who we aptly named Mr. B. For Illumina sequencing, reads were generated at Duke University (16×), Illumina UK (54×), and BGI (219×) using Illumina’s TruSeq [1] version2 or version3 chemistries (Table 1 and GigaDB [2]). The version3 chemistry reads through GC-rich regions, which are often found in promoters, more evenly than does version2 [3]. The insert sizes for the BGI libraries ranged from 220 bp to 40 Kbp, and the insert sizes for the Duke libraries ranged from 400–600 bp, in order to assist assemblies. Fragment sizes for the mate pair libraries, based on genome mapping, and the per base sequence quality distribution for the libraries are shown in GigaDB [2]. The Duke University Illumina libraries were sequenced at two different cluster densities: 8× coverage reads at the normal 420 k clusters/mm density and 8× coverage at a lower 350 k clusters/mm. The lower cluster density was used to increase the number of GC-rich regions sequenced. For PacBio sequencing, 6.76 Gbp (~5.5× coverage) of PacBio RS reads [4] were generated at Pacific Biosciences from two insert size libraries (7.5 K bp at 1.93× and 13 Kbp at 3.56×; PacBio reads error-corrected with Illumina can be downloaded from the supplementary webpage associated with [5]). With all reads combined, the total coverage exceeds 300× (assuming a haploid genome size of 1.23 Gbp) (Table 1), perhaps making Mr. B one of the most sequenced individual vertebrate animals as of to date. The read length distributions of these different types of reads are shown in Figure 1.

Bottom Line: The sequence reads for this project were in part generated and used for both the Assemblathon 2 competition and the first de novo assembly of a giga-scale vertebrate genome utilizing PacBio single-molecule sequencing.Across several quality metrics, these budgerigar assemblies are comparable to or better than the chicken and zebra finch genome assemblies built from traditional Sanger sequencing reads, and are sufficient to analyze regions that are difficult to sequence and assemble, including those not yet assembled in prior bird genomes, and promoter regions of genes differentially regulated in vocal learning brain regions.This work provides valuable data and material for genome technology development and for investigating the genomics of complex behavioral traits.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Neurobiology, Duke University Medical Center, Durham, NC 27710, USA.

ABSTRACT

Background: Parrots belong to a group of behaviorally advanced vertebrates and have an advanced ability of vocal learning relative to other vocal-learning birds. They can imitate human speech, synchronize their body movements to a rhythmic beat, and understand complex concepts of referential meaning to sounds. However, little is known about the genetics of these traits. Elucidating the genetic bases would require whole genome sequencing and a robust assembly of a parrot genome.

Findings: We present a genomic resource for the budgerigar, an Australian Parakeet (Melopsittacus undulatus) -- the most widely studied parrot species in neuroscience and behavior. We present genomic sequence data that includes over 300× raw read coverage from multiple sequencing technologies and chromosome optical maps from a single male animal. The reads and optical maps were used to create three hybrid assemblies representing some of the largest genomic scaffolds to date for a bird; two of which were annotated based on similarities to reference sets of non-redundant human, zebra finch and chicken proteins, and budgerigar transcriptome sequence assemblies. The sequence reads for this project were in part generated and used for both the Assemblathon 2 competition and the first de novo assembly of a giga-scale vertebrate genome utilizing PacBio single-molecule sequencing.

Conclusions: Across several quality metrics, these budgerigar assemblies are comparable to or better than the chicken and zebra finch genome assemblies built from traditional Sanger sequencing reads, and are sufficient to analyze regions that are difficult to sequence and assemble, including those not yet assembled in prior bird genomes, and promoter regions of genes differentially regulated in vocal learning brain regions. This work provides valuable data and material for genome technology development and for investigating the genomics of complex behavioral traits.

No MeSH data available.


Related in: MedlinePlus