![]() It is imperative to understand that prediction and annotation of non-protein-coding genes, Untranslated Regions (UTR), and tRNA are as vital as protein-coding genes to determine the overall genetic constitution of the assembled genome. This step entails the prediction of all the genes present in the assembled genome and to provide efficient functional annotation to these genes from the data available in diverse public repositories such as Protein Family ( PFAM), SuperFamily, Conserved Domain Database ( CDD), TIGRFAM, PROSITE, CATH, SCOP, and other protein domain databases. Figure 6: Transcripts annotation completeness comparison based on BUSCO results between Pipelines (GeMoMa vs MAKER) and within MAKER training.After genome assembly (covered in my previous blog) comes the vital step of gene prediction and annotation. Figure 5: Protein annotation completeness comparison based on BUSCO results between Pipelines (GeMoMa vs MAKER) and within MAKER training. Changes of annotation numbers during MAKER training. Figure 4: Summary of predictions between annotation pipeline GeMoMa and MAKER. An increasing accuracy during the training is noticeable (more annotations covered by RNA data with lower AED). Figure 3: Annotation Edit Distances (AED) of all performed MAKER rounds (within MAKER training). ![]() Transcriptsĭownload unique transcripts fasta from (15,518 sequences): P.cal transcript sequences.fasta Proteinsĭownload proteins according to unique transcripts (15,518 sequences): P.cal protein sequences.fasta Annotationĭownload structural annotation of unique transcripts gff: P.cal final predictions.gff Discussion Figure 2: Comparison of Annotation completeness between relative species. MRNA in the gff file refers to the gene and its region in the genome. Resultsĭescription: Ending of the protein ID (with RB, RC, RD etc.) refers to the isoforms of the protein coding gene. Gene finders are used as Augusts and SNAP. The RepeatMasker step was skipped, because it was performed earlier with a customized repeat database. The GHMM from the Augustus training were used to train MAKER in the fourth and last round. The third MAKER round was done by using a BUSCO trained Augusts reference model based on single copy ortholgous genes from Hymenoptera clades in the P. This was done in order to provide a reduction of false positive protein predictions. Resulting MAKER predictions from the previous run with an AED smaller that 0.25 were handed to SNAP for generating more accurate gene models to train MAKER in the next round. For the first run of MAKER, the Nasonia reference model for Augustus detection was used. The data from GeMoMa as well as the data from relatives were handed to the pipeline (see figure 2). MAKER can be trained by itself and by other programs like Augustus or SNAP. The AED is a number between 0 and 1, where 0 means the entire gene is covered by RNA matching regions. The Annotation Edit Distance (AED) was developed (Eilbeck et al., 2009 Holt and Yandell, 2011 Yandell and Ence, 2012) to measure the evidence of the annotations coming from MAKER. Additionally, MAKER uses tRNAscan ffor detection of tRNA. ![]() SNAP deals with Hidden Markoc Models (HMMs), as does Augustus, but this program uses these models to calculate similarities of intron length probabilities. ![]() ![]() Augusts predicts genes in eukaryotic genomic sequences by using the Generalized Hidden Markov Model (GHMM) to align Expressed Sequence Tags (ESTs) and proteins to the genome. MAKER makes use of gene prediction software, Augustus and Semi-HMM-based Nucleic Acid Parser (SNAP), as gene finders. The repeat annotation step was skipped because it was performed earlier with a custom designed repeat database (see Repeat masking section ). The MAKER algorithm uses different prediction programs (see figure 1) for mask repetitive elements in the genome (RepeatMasker), aligns proteins and RNA evidence to the genome assembly (BLAST), includes data from relatives, and identifies splice sites (Exonerate). MAKER 2 is a genome annotation pipeline for smaller eukaryotic and prokaryotic genomes. Summary of predictions from the MAKER pipeline. ![]()
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