Dataset Information

Next-generation pyrosequencing of gonad transcriptomes in the polyploid lake sturgeon (Acipenser fulvescens): the relative merits of normalization and rarefaction in gene discovery.

ABSTRACT: BACKGROUND:Next-generation sequencing technologies have been applied most often to model organisms or species closely related to a model. However, these methods have the potential to be valuable in many wild organisms, including those of conservation concern. We used Roche 454 pyrosequencing to characterize gene expression in polyploid lake sturgeon (Acipenser fulvescens) gonads. RESULTS:Titration runs on a Roche 454 GS-FLX produced more than 47,000 sequencing reads. These reads represented 20,741 unique sequences that passed quality control (mean length = 186 bp). These were assembled into 1,831 contigs (mean contig depth = 4.1 sequences). Over 4,000 sequencing reads (approximately 19%) were assigned gene ontologies, mostly to protein, RNA, and ion binding. A total of 877 candidate SNPs were identified from > 50 different genes. We employed an analytical approach from theoretical ecology (rarefaction) to evaluate depth of sequencing coverage relative to gene discovery. We also considered the relative merits of normalized versus native cDNA libraries when using next-generation sequencing platforms. Not surprisingly, fewer genes from the normalized libraries were rRNA subunits. Rarefaction suggests that normalization has little influence on the efficiency of gene discovery, at least when working with thousands of reads from a single tissue type. CONCLUSION:Our data indicate that titration runs on 454 sequencers can characterize thousands of expressed sequence tags which can be used to identify SNPs, gene ontologies, and levels of gene expression in species of conservation concern. We anticipate that rarefaction will be useful in evaluations of gene discovery and that next-generation sequencing technologies hold great potential for the study of other non-model organisms.

SUBMITTER: Hale MC

PROVIDER: S-EPMC2688523 | biostudies-literature | 2009 Apr

REPOSITORIES: biostudies-literature

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Next-generation pyrosequencing of gonad transcriptomes in the polyploid lake sturgeon (Acipenser fulvescens): the relative merits of normalization and rarefaction in gene discovery.

Hale Matthew C MC McCormick Cory R CR Jackson James R JR Dewoody J Andrew JA

BMC genomics 20090429

<h4>Background</h4>Next-generation sequencing technologies have been applied most often to model organisms or species closely related to a model. However, these methods have the potential to be valuable in many wild organisms, including those of conservation concern. We used Roche 454 pyrosequencing to characterize gene expression in polyploid lake sturgeon (Acipenser fulvescens) gonads.<h4>Results</h4>Titration runs on a Roche 454 GS-FLX produced more than 47,000 sequencing reads. These reads r ...[more]

PMID: 19402907

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Project description:In this study, the effects of different feeding frequencies on the growth and the expression of genes in the GH/IGF axis were assessed in juvenile Chinese sturgeon. The newly hatched Chinese sturgeons were bred for 38 days at three different feeding frequencies groups (feeding frequency of two times a day, TWD; three times a day, THD; and four times a day, FOD), and the expression levels of the GH/IGF axis responses to feeding frequency were determined by quantitative real-time PCR. In addition, the full-length of the Coding Sequences of IGF I and IGF II genes (489-bp and 660-bp, respectively), were cloned and analyzed from Chinese sturgeon the first time. Multiple sequence alignments of IGFs revealed that Chinese sturgeon are high sequence identity to IGFs from other species. The phylogenetic relationships based on the IGF I and IGF II amino acid sequences were consistent with the traditional classification. After 38 days of growth, the three different feeding frequencies groups of Chinese sturgeon had no significant difference of body length, body weight, specific growth rate, the survival rate, the rate of weight gain and the condition factor. However, the relative expression of Chinese sturgeon GH in the pituitary decreased with increasing feeding frequency. The relative expression of Chinese sturgeon GHR in liver and skeletal muscle was deceased with increasing feeding frequency, while the relative expression of GHR in stomach and intestines at THD group was significantly higher than that of at TWD group and FOD group (p?<?0.05). The relative expression of Chinese sturgeon IGF I in liver increased significantly with increasing feeding frequency (p?<?0.05). The relative expression of IGF I in stomach and skeletal muscle was similar at the three groups. The relative expression of IGF I in intestines was significantly higher at FOD group than at TWD group and THD group (p?<?0.05). The relative expression of Chinese sturgeon IGF II in liver at TWD group was significantly higher than that at THD group and FOD group (p?<?0.05). However, the relative expression of IGF II in stomach, intestines and skeletal muscle at THD group was higher than that at TWD group and FOD group. Based on these previous studies that liver IGF I is regarded as a biomarker of growth performance, this result suggested that the juvenile Chinese sturgeon is better for growth when feeding four times daily compared to twice and thrice daily.

Project description:BackgroundEvolution of sturgeons and paddlefishes (order Acipenseriformes) is inherently connected with polyploidization events which resulted in differentiation of ploidy levels and chromosome numbers of present acipenseriform species. Moreover, allopolyploidization as well as autopolyploidization seems to be an ongoing process in these fishes and individuals with abnormal ploidy levels were occasionally observed within sturgeon populations. Here, we reported occurrence of Siberian sturgeon (Acipenser baerii) male with abnormal ploidy level for this species, accessed its ploidy level and chromosome number and investigate its potential sterility or fertility in comparison with normal individuals of sterlet (A. ruthenus), Russian sturgeon (A. gueldenstaedtii) and Siberian sturgeon (A. baerii).ResultsAcipenser ruthenus possessed 120 chromosomes, exhibiting recent diploidy (2n), A. gueldenstaedtii and A. baerii had ~245 chromosomes representing recent tetraploidy (4n), and A. baerii male with abnormal ploidy level had ~ 368 chromosomes, indicating recent hexaploidy (6n). Genealogy assessed from the mtDNA control region did not reveal genome markers of other sturgeon species and this individual was supposed to originate from spontaneous 1.5 fold increment in number of chromosome sets with respect to the number most frequently found in nature for this species. Following hormone stimulation, the spontaneous hexaploid male produced normal sperm with ability for fertilization. Fertilization of A. baerii and A. gueldenstaedtii ova from normal 4n level females with sperm of the hexaploid male produced viable, non-malformed pentaploid (5n) progeny with a ploidy level intermediate to those of the parents.ConclusionThis study firstly described occurrence of hexaploid individual of A. baerii and confirmed its autopolyploid origin. In addition to that, the first detailed evidence about fertility of spontaneous hexaploid sturgeon was provided. If 1.5 fold increment in number of chromosome sets occurring in diploids, resulted triploids possess odd number of chromosome sets causing their sterility or subfertility due to interference of gametogenesis. In contrast, 1.5 fold increment in number of chromosome sets in naturally tetraploid A. baerii resulted in even number of chromosome sets and therefore in fertility of the hexaploid specimen under study.

Dataset Information

Next-generation pyrosequencing of gonad transcriptomes in the polyploid lake sturgeon (Acipenser fulvescens): the relative merits of normalization and rarefaction in gene discovery.

Publications

Next-generation pyrosequencing of gonad transcriptomes in the polyploid lake sturgeon (Acipenser fulvescens): the relative merits of normalization and rarefaction in gene discovery.

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