Systematic Analysis and Expression Profiles of the 4-Coumarate: CoA Ligase (4CL) Gene Family in Pomegranate (Punica granatum L.)
Abstract
:1. Introduction
2. Results
2.1. Identification of Pg4CL Family Genes and Gene Cloning of Pg001327.1
2.2. Analysis of Physical and Chemical Properties
2.3. Phylogenetic Analysis and Classification of the Pg4CL Gene Family
2.4. Analysis of Conserved Motifs and Gene Structures of Pg4CL Gene Family
2.5. Cis-Acting Elements in the Promoter Region of Pg4CLs
2.6. Protein Interaction Networks of Pg4CL Gene Family
2.7. Expression Analysis of Pg4CL Genes with RNA-Seq
2.8. qRT-PCR Analysis of Pg4CLs during Fruit Development in Pomegranate
3. Discussion
4. Materials and Methods
4.1. Genome and Transcriptome Data Sources
4.2. Identification of Pg4CL Family Genes
4.3. Gene Cloning of Pg001327.1
4.4. Virtual Prediction of Physical and Chemical Properties
4.5. Multiple Sequences Alignment and Phylogenetic Analysis
4.6. Analysis of Gene Structure and Protein Conserved Motifs
4.7. Analysis of Cis-Acting Elements and Protein–Protein Interaction Networks
4.8. RNA-Seq Analysis of 4CL Gene Family in Pomegranate
4.9. Expression Patterns by Quantitative Real-Time PCR (qRT-PCR)
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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Gene Name | Gene ID | Location | CDS (bp) | Exon No. | AA | pI | Instability Index | GRAVY | Signal Peptide | Protein Localization |
---|---|---|---|---|---|---|---|---|---|---|
Pg4CL1 | Pg000024.1 | scaffold1:6623488:6634629 | 2343 | 14 | 780 | 8.81 | 47.22 | 0.059 | No | Plasma membrane |
Pg4CL2 | Pg001327.1 | scaffold10:76276:83034 | 1887 | 6 | 628 | 5.83 | 46.64 | 0.066 | No | Cytosol |
Pg4CL3 | Pg004702.1 | scaffold124:253916:256949 | 1674 | 6 | 557 | 6.55 | 39.93 | 0.159 | No | Plasma membrane |
Pg4CL4 | Pg005072.1 | scaffold13:4695163:4697720 | 1755 | 6 | 584 | 8.09 | 49.09 | −0.057 | No | Extracellular |
Pg4CL5 | Pg007092.1 | scaffold15:2592574:2599061 | 1635 | 6 | 544 | 5.65 | 36.60 | −0.034 | No | Extracellular |
Pg4CL6 | Pg007674.1 | scaffold152:355399:358070 | 1629 | 6 | 542 | 8.64 | 26.44 | 0.044 | No | Plasma membrane |
Pg4CL7 | Pg013255.1 | scaffold22:4058452:4060831 | 1638 | 7 | 545 | 7.65 | 38.71 | −0.023 | No | Plasma membrane |
Pg4CL8 | Pg015184.1 | scaffold27:1309358:1311530 | 1722 | 4 | 573 | 5.49 | 40.47 | 0.024 | No | Plasma membrane |
Pg4CL9 | Pg016048.1 | scaffold3:3588368:3591221 | 1644 | 6 | 547 | 5.64 | 35.48 | 0.120 | No | Cytosol |
Pg4CL10 | Pg016524.1 | scaffold3:3410537:3413121 | 1659 | 6 | 552 | 8.45 | 42.89 | 0.153 | No | Peroxisome |
Pg4CL11 | Pg018200.1 | scaffold35:541618:543985 | 1656 | 6 | 551 | 8.7 | 41.33 | 0.037 | No | Plasma membrane |
Pg4CL12 | Pg029185.1 | scaffold85:64794:69120 | 1731 | 6 | 576 | 8.91 | 44.18 | −0.010 | No | Plasma membrane |
Motif Code | Sequences | E_Value | Characteristic Domain |
---|---|---|---|
Motif 1 | DGWLHTGDLCYFDDEGFLFIVDRJKELIKYKGYQVAPAELEALLLSHPEI | 6.2 × 10−341 | AMP-binding enzyme |
Motif 2 | GRLVPNMEAKIVDPETGAALPPNQSGELWLRGPTIMKGYLNBPEATAETI | 5.20 × 10−242 | AMP-binding enzyme |
Motif 3 | DAAVIPYPDEEAGEIPMAFVVRKPGSSITEEDVIDFVAKQVAPYKKIRRV | 7.40 × 10−236 | No |
Motif 4 | KGDVVLILLPNSJHFPIIYLSILSLGAVITTANPLSTPSEIAKQVRDSKP | 9.60 × 10−162 | AMP-binding enzyme |
Motif 5 | VRVDQDDTAAJLYSSGTTGTSKGVVLTHRNLIA | 1.90 × 10−160 | AMP-binding enzyme |
Motif 6 | KSSLTDKYDLSSLRQVGSGAAPLGKEVAEQFRKKFPHVELLQGYGMTEST | 2.90 × 10−160 | AMP-binding enzyme |
Motif 7 | LRAGETVVVMQKFEFEAMLRAIEKYRVTYLPVVPPIILALV | 4.60 × 10−102 | No |
Motif 8 | FIESIPKSPAGKILRRELIKK | 2.30 × 10−81 | No |
Motif 9 | NVFLCVLPLFHIYGLAVIILG | 2.70 × 10−53 | No |
Motif 10 | SGFCSKTGIFHSLRPPVPLPP | 3.90 × 10−32 | No |
NO. | Accession NO. | Cultivars | Sample Type | Library | Platform | Reference |
---|---|---|---|---|---|---|
P1 | SRR5279396 | ‘Dabenzi’ | root | Paired end | Illumina HiSeq 4000 | [26] |
P2 | SRR5279397 | ‘Dabenzi’ | leaf | Paired end | Illumina HiSeq 4000 | [26] |
P3 | SRR5279395 | ‘Dabenzi’ | flower | Paired end | Illumina HiSeq 4000 | [26] |
P4 | SRR5279391 | ‘Dabenzi’ | Pericarp | Paired end | Illumina HiSeq 4000 | [26] |
P5 | SRR5279388 | ‘Dabenzi’ | Exocarp | Paired end | Illumina HiSeq 4000 | [26] |
P6 | SRR5279394 | ‘Dabenzi’ | Pericarp | Paired end | Illumina HiSeq 4000 | [26] |
P7 | SRR5678820 | ‘Tunisia’ | Pericarp | Paired end | Illumina HiSeq 4000 | [26] |
P8 | SRR5678819 | ‘Baiyushizi’ | Pericarp | Paired end | Illumina HiSeq 4000 | [26] |
P9 | SRR080723 | ‘Wonderful’ | Pericarp | Paired end | Illumina HiSeq 2000 | [52] |
P10 | SRR1055290 | ‘Nana’ | Mixed samples of the root, | Single end | 454 GS FLX Titanium | [53] |
leaves, flowers, and fruit | ||||||
P11 | SRR1054190 | ‘Black127’ | Mixed samples of the root, | Single end | 454 GS FLX Titanium | [53] |
leaves, flowers, and fruit |
Gene | Primer Sequence (5′-3′) |
---|---|
Pg4CL1 | F: GGATGGCTGGTTGAGGACAG; R: GCAGCATCTGCAATTTCGGG |
Pg4CL2 | F: CAGTGCTCTCGATGTGCCTA; R: TCATAATCTGTTGGCCTCGAA |
Pg4CL3 | F: AGAATGTGGGGAGGTCCCAG; R: CCTGCTGGAGACTTCGGTATC |
Pg4CL4 | F: AGAAGCAGGACAGATCCCGA; R: CTTCCCTGCCGGAGATTTCG |
Pg4CL5 | F: GCATTCGTCATCCGATCCAA; R: CGGATGCCAATTTCGCTCT |
Pg4CL6 | F: GCTTTCAGATTGCTCCTGCG; R: ATTCTGCACGTCCTCTTCGG |
Pg4CL7 | F: CCGGAGACCTCGGGTATTTC; R: TCGGGAAACGGAATGACCAC |
Pg4CL8 | F: AGTTGTACCGGTCGAGGATG; R: CTTGTAAGGCGCAACCTGAG |
Pg4CL9 | F: CGAATTGTGTGTCCGAAGCC; R: ACGACAAAGACGTCCCCATC |
Pg4CL10 | F: TACTCCCATCCCGACATTGC; R: GTCGTTTCACCACGAAAGCC |
Pg4CL11 | F: TCCGAACGGGAGATCTTTGC; R: TCGGGAAACGGGATAACAGC |
Pg4CL12 | F: GCAGATGCCGCTGTTATTCC; R: TTCCCTGCCGGAGATTTCG |
PgActin | F: AGTCCTCTTCCAGCCATCTC; R: CACTGAGCACAATGTTTCCA |
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Wang, Y.; Guo, L.; Zhao, Y.; Zhao, X.; Yuan, Z. Systematic Analysis and Expression Profiles of the 4-Coumarate: CoA Ligase (4CL) Gene Family in Pomegranate (Punica granatum L.). Int. J. Mol. Sci. 2022, 23, 3509. https://doi.org/10.3390/ijms23073509
Wang Y, Guo L, Zhao Y, Zhao X, Yuan Z. Systematic Analysis and Expression Profiles of the 4-Coumarate: CoA Ligase (4CL) Gene Family in Pomegranate (Punica granatum L.). International Journal of Molecular Sciences. 2022; 23(7):3509. https://doi.org/10.3390/ijms23073509
Chicago/Turabian StyleWang, Yuying, Linhui Guo, Yujie Zhao, Xueqing Zhao, and Zhaohe Yuan. 2022. "Systematic Analysis and Expression Profiles of the 4-Coumarate: CoA Ligase (4CL) Gene Family in Pomegranate (Punica granatum L.)" International Journal of Molecular Sciences 23, no. 7: 3509. https://doi.org/10.3390/ijms23073509