Cloning and expression analysis of CCoAOMT gene in fruit of ‘Dangshansuli'
Bibliographic record
Abstract
[Objectives]Lignin is widely distributed in plant cell walls and confers strength,rigidity and impermeability,which are the principal components of stone cells in pear. Caffeoyl-CoA O-methyltransferase( CCoAOMT) plays an important role in the biosynthesis of lignin,and the 3-O-methylation of caffeoyl CoA leading to the formation of feruloyl Co A is catalyzed by the enzyme CCoAOMT.[Methods]In order to investigate the function of CCoAOMT in the synthesis of pear stone cells,15-year-old trees of Pyrus bretschneideri‘Dangshansuli'were used in this study,and we cloned Pb CCoAOMT gene and analyzed its biological information,and studied the gene expression at different stages during pear fruit development. These results contributed to providing some theoretical reference and practical basis to improve the quality of pear. Comparing CCoAOMT gene sequences associated with lignification obtained from from the NCBI database with pear genome database,the Pb CCoAOMT gene cDNA sequence involved in lignin biosynthesis was cloned by RT-PCR techniques. Gene structure analysis relied on GSDS. Sequence alignments and homology sequence searchers in databases were carried out using NCBI BLAST webpage. Deduced amino acid sequences were obtained by Primer Premier 5.0 software. Phylogenetic relationships among sequences were performed with MEGA 6.06. The molecular structure and physicochemical properties of proteins were analyzed by Prot Param. Hydrophobic analysis of deduced protein sequences was demonstrated by means of Prot Scale. Signal peptide of deduced protein sequences was predicted by Bio XM 2. 6. The transmembrane domain of deduced protein sequences was predicted by TMPRED. Secondary and tertiary structures of deduced protein sequences were aligned by SOPMA and SWISS-MODEL separately. For gene expression analysis,total genomic RNA was isolated from fruit samples collected at different stages during pearfruit development according to the CTAB method and c DNAs were generated following the manufacturers' recommendations to a final volume of 20 μL,gene quantifications were performed according to the manufacturers' instructions and Pyrus EFα1( EFα1,accession No.: AY338250) was used as internal gene to evaluate the RT-qPCR assays. [Results]The full-length cDNA of Pb CCoAOMT( Gen Bank accession No.: KJ577544) in pear was cloned. The sequence consists of 1 133 bp with an open reading frame of 744 bp,which encodes a polypeptide of 247 amino acids residues. Sequence alignment and phylogenetic analysis showed that CCoAOMT is a member of O-methyltransferases family and shared high similarity with CCoAOMT proteins from other species,especially with the highest similarity of 98.79% and phylogenetic relationship compared with the loquat. Real-time PCR results showed that the change of Pb CCoAOMT expression level was similar to that of stone cell content during fruit development,which showed a rise-down tendency.[Conclusions]Pb CCoAOMT gene was firstly isolated and characterized from pear,which may be involved in metabolism of lignin.
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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 | 0.001 |
| 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".