Identification and Characterization of Novel Genes from Blastocyst-Stage Porcine Embryos Using 454 Sequencing.
Bibliographic record
Abstract
During the pre-implantation period of embryonic development, the mammalian embryo exhibits dramatic morphological changes and many key developmental events take place. Due to physiological similarities between pig and human, there is strong interest in producing large quantities of pre-implantation embryos through assisted reproductive technologies for biomedical and biological research purposes. The developmental stage at which porcine embryos are most commonly manipulated is the blastocyst, and the quality of the embryos utilized is critical to the success of reproductive technologies such as embryo transfer. Despite the importance of this critical developmental period, our ability to determine early embryonic quality based on morphological criteria is limited. As an alternative, characterization of the early porcine embryo transcriptome could identify gene markers of embryonic quality. 454 sequencing is a powerful ultra-high-throughput method of gene expression profiling which is able to characterize the expression of both known and novel genes. In this study, 454 sequencing was performed on a cDNA library constructed from 15 in vivo-derived porcine blastocysts using a whole transcriptome amplification method (Ribo-SPIA, NuGen) in order to identify and characterize novel genes from blastocyst-stage embryos. A total of 19,438 reads were generated from a 1/8 plate run using GS FLX Titanium reagent kit (454 Technology, Roche), resulting in 18,338 quality ESTs (expression sequence tags) after filtering using 454 Replicate Filter (http://microbiomes.msu.edu/replicates/), of which approximately 1.1% (202 ESTs) were identified as unknown ESTs with significant matches in the porcine EST database (http://www.ncbi.nlm.nih.gov/genome/seq/BlastGen/BlastGen.cgi?taxid=9823). A consensus sequence of those porcine EST matches was identified for each unknown EST query using the Seqman module of DNAstar software (http://www.dnastar.com). The consensus sequences were further compared with the NCBI nucleotide database using the NCBI BLAST tool (http://blast.ncbi.nlm.nih.gov/Blast.cgi), and 41 of the sequences had significant matches with porcine refseq RNA records. An additional 156 sequences were identified as novel genes and were further analyzed using the Database for Annotation, Visualization and Integrated Discovery (DAVID) functional annotation tool (http://david.abcc.ncifcrf.gov/tools.jsp). The results indicate that many of these newly identified genes are involved with pathways important to embryonic development, such as pathways related to differentiation, apoptosis, cell survival and proliferation. The findings from this study increase our understanding of the molecular mechanisms involved in early porcine embryonic development, and may help to identify candidate gene markers of early porcine embryo quality. This research was supported by the Natural Sciences and Engineering Research Council (NSERC) and is a part of the activities of the EmbryoGENE NSERC Strategic Research Network. (poster)
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How this classification was reachedexpand
Full frame distilled prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. Learned from the 10,348 direct Codex labels and 10,348 direct Gemma labels. Candidate is the union of thresholded teacher heads; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels or direct frontier model labels.
Codex and Gemma teacher scores by category
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.000 |
| Insufficient payload (model declined to judge) | 0.000 | 0.000 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one teacher head, not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".