Transcriptome Analysis of In Vitro- and In Vivo-Produced Preimplantation Porcine Embryos by 454 Pyrosequencing.
Bibliographic record
Abstract
Currently, our understanding of the molecular events during the development of porcine preimplantation embryos is limited. Increased knowledge in this area will contribute to our understanding of basic reproductive biology, will allow for identification of molecular markers related to embryonic quality, and will facilitate improvement to in vitro embryo production and culture conditions. The objective of this study was to identify transcripts related to the development of in vitro- (IVT) and in vivo (IVV)-produced porcine embryos using the 454-sequencing platform. Two normalized cDNA libraries were generated from pools of IVT and IVV embryos at different developmental stages (oocyte, 2-cell, 4-cell, 8-cell, morula and blastocyst). In order to facilitate the identification of rare transcripts, TRIMMER kit (Evrogen, Moscow, Russia) was used to decrease the prevalence of abundant transcripts and increase the efficiency of 454 sequencing for discovery of rare copies. A preliminary sequence analysis of the IVT and IVV cDNA libraries from a 1/8 plate run on the 454-Genome Sequencer FLX system (Roche) using the Titanium reagent kit yielded 94,628 and 82,582 ESTs, respectively. NextGENe (SoftGenetics LLC) software was used to identify expressed genes from a variety of porcine sequence databases as references. These reference sequence collections were downloaded from Ensembl: Sus_scrofa.Sscrofa9.56.cdna.abinitio.fa, Sus_scrofa.Sscrofa9.56.cdna.all.fa, and Sus_scrofa.Sscrofa9.56.dna.chromosome.*.fa; and from NCBI: rna.gbk. The collection of chromosomes were indexed using the ‘Build Index for WGA’ (whole genome alignment) tool of NextGENe. An average of 20% of expressed genes matched to the above references in ESTs obtained from both normalized cDNA libraries. Differential expression levels for IVT and IVV embryos were compared using the RPKM (reads per kilobase of exon model per million mapped reads) method and mapped to two databases (the NCBI rna.gbk genome and a combined Ensembl abinitio and cdna-all reference). Some commonly expressed genes were identified, and also, unique genes expressed either in IVT or IVV embryos. The remaining uncharacterized transcripts had significant matches against the EST porcine database. We further annotated the transcripts using the non-porcine mammalian protein database and Gene Ontology terms. This newly found EST resource will be valuable for embryo specific microarray development and for comparative mammalian genome analysis. 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".