A Combinatorial Single-Molecule Real-Time and Illumina Sequencing Analysis of Postembryonic Gene Expression in the Asian Citrus Psyllid Diaphorina citri
Abstract
:Simple Summary
Abstract
1. Introduction
2. Materials and Methods
2.1. D. citri Rearing, Sample Collection, and Assay of Weight and Length
2.2. RNA Isolation and cDNA Synthesis
2.3. cDNA Library Construction and Sequencing
2.4. Functional Annotation of Full-Length, Non-Redundant Transcripts
2.5. Alternative Splicing (AS) Event, Simple Sequence Repeat (SSR), and Open Reading Frame (ORF) Prediction
2.6. Long Noncoding RNA (LncRNA) Prediction and Transcription Factor (TF) Analysis
2.7. Identification of DEGs and Functional Enrichment Analysis
2.8. dsRNA Synthesis and Microinjection
2.9. Analysis of Expression Levels of DEGs in Different Samples by RT-qPCR
2.10. Statistical Analysis
3. Results
3.1. Detection of Weight and Length of D. citri Fifth-Instar Nymphs and Adults
3.2. PacBio and Illumina Sequencing of D. citri Nymphs and Adults
3.3. Functional Annotation of New Transcripts
3.4. Analysis of SSRs, ORFs, and LncRNAs
3.5. Prediction of TFs, AS Events, and APA
3.6. Identification of Differentially Expressed Genes (DEGs) and Functional Enrichment Analysis
3.7. Validation of DEGs by Reverse Transcription–Quantitative PCR (RT-qPCR)
3.8. Analysis of Spatiotemporal Expression Patterns of DEGs Involved in the Hippo Signaling Pathway
3.9. Silencing of DcYki by RNAi Affects the Development of D. citri
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Data Availability Statement
Conflicts of Interest
References
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Data Statistics | Nymph (Number) | Adult (Number) |
---|---|---|
SMRT cells | 3 | 1 |
cDNA size | 1–6 K | 1–6 K |
Data size (G) | 20.17 | 23.97 |
Number of CCSs | 306,224 | 347,641 |
Read bases of CCSs | 806,848,990 | 638,839,227 |
Mean read length of CCSs | 2634 | 1837 |
Mean number of passes | 28 | 44 |
Number of undesired primer reads | 77,099 | 78,527 |
Number of filtered short reads | 49 | 89 |
Number of full-length, non-chimeric reads | 218,559 | 256,717 |
Full-length, non-chimeric percentage (FLNC %) | 71.37% | 73.85% |
Number of consensus isoforms | 16,119 | 20,157 |
Average consensus isoform read length | 2559 | 1778 |
Number of polished, high-quality isoforms | 15,992 | 20,048 |
Number of polished, low-quality isoforms | 126 | 105 |
Number of full-length, non-redundant transcripts | 4307 | 6334 |
Percentage of polished, high-quality isoforms | 99.22% | 99.48% |
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Zhang, Q.; Zhang, C.; Zhong, H.; He, Q.; Xia, Z.-Y.; Hu, Y.; Liao, Y.-X.; Yi, L.; Lu, Z.-J.; Yu, H.-Z. A Combinatorial Single-Molecule Real-Time and Illumina Sequencing Analysis of Postembryonic Gene Expression in the Asian Citrus Psyllid Diaphorina citri. Insects 2024, 15, 391. https://doi.org/10.3390/insects15060391
Zhang Q, Zhang C, Zhong H, He Q, Xia Z-Y, Hu Y, Liao Y-X, Yi L, Lu Z-J, Yu H-Z. A Combinatorial Single-Molecule Real-Time and Illumina Sequencing Analysis of Postembryonic Gene Expression in the Asian Citrus Psyllid Diaphorina citri. Insects. 2024; 15(6):391. https://doi.org/10.3390/insects15060391
Chicago/Turabian StyleZhang, Qin, Can Zhang, Hong Zhong, Qing He, Zhao-Ying Xia, Yu Hu, Yu-Xin Liao, Long Yi, Zhan-Jun Lu, and Hai-Zhong Yu. 2024. "A Combinatorial Single-Molecule Real-Time and Illumina Sequencing Analysis of Postembryonic Gene Expression in the Asian Citrus Psyllid Diaphorina citri" Insects 15, no. 6: 391. https://doi.org/10.3390/insects15060391