Single-Molecule Long-Read Sequencing of Avocado Generates Microsatellite Markers for Analyzing the Genetic Diversity in Avocado Germplasm
Abstract
:1. Introduction
2. Materials and Methods
2.1. Sample Collection, DNA Extraction, and RNA Extraction
2.2. PacBiocDNA Library Construction and Sequencing
2.3. IlluminacDNA Library Construction and Sequencing
2.4. Quality Filtering and Correction of PacBio Long-Reads
2.5. Functional Annotation
2.6. Mining of EST-SSR Markers
2.7. Analyses of Detected Coding Sequences, Transcription Factors, and Long Non-Coding RNA Features
2.8. Assignment of the Native Avocado Accessions with an Unknown Race
2.9. Identification of EST-SSR Markers
2.10. Data Analysis
3. Results
3.1. General Properties and Functional Annotations Based on Public Databases of Single-Molecule Long-Reads
3.2. Predictions of ORFs, TFs, and lncRNAs
3.3. Frequency and Distribution of Various Types of EST-SSR Loci
3.4. Development of Polymorphic EST-SSR Markers, Analysis of Genetic Diversity, and KASP genotyping
3.5. Analyses of Genetic Relationships Based on Polymorphic EST-SSRs from SMRT Sequencing Data
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
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Source | Number |
---|---|
Total number of sequences examined | 75,956 |
Total size of examined sequences (bp) | 170,959,769 |
Total number of identified SSRs | 76,777 |
Number of SSR containing sequences | 42,096 |
Number of sequences containing more than 1 SSR | 19,825 |
Number of SSRs present in compound formation | 12,675 |
SSR Motif Length | Repeat Unit Number | ||||||||
---|---|---|---|---|---|---|---|---|---|
5 | 6 | 7 | 8 | 9 | 10 | >10 | Total | % | |
Mono- | - | - | - | - | - | 8951 | 35,849 | 44,800 | 58.35 |
Di- | - | 4017 | 2773 | 2489 | 2013 | 1547 | 6064 | 18,903 | 24.62 |
Tri- | 6129 | 2838 | 1237 | 720 | 403 | 193 | 204 | 11,724 | 15.27 |
Tetra- | 541 | 175 | 41 | 21 | 8 | - | 2 | 788 | 0.01 |
Penta- | 172 | 67 | 1 | 1 | - | - | - | 241 | 0.00 |
Hexa- | 228 | 72 | 15 | 3 | 2 | - | 1 | 321 | 0.00 |
Total | 7070 | 7169 | 4067 | 3234 | 2426 | 10,691 | 42,120 | 76,777 | |
% | 9.21 | 9.34 | 5.30 | 4.21 | 3.16 | 13.92 | 45.14 |
Marker Name | Transcript ID | Na 1 | Ne 2 | Ho 3 | He 4 | PIC 5 |
---|---|---|---|---|---|---|
Pa-eSSR-17 | F01_cb7709_c10/f1p0/2063 | 8 | 3.02 | 0.61 | 0.67 | 0.62 |
Pa-eSSR-18 | F01_cb7876_c2/f1p0/2226 | 10 | 3.09 | 0.61 | 0.68 | 0.65 |
Pa-eSSR-19 | F01_cb1803_c26/f1p0/2838 | 6 | 2.04 | 0.63 | 0.51 | 0.46 |
Pa-eSSR-20 | F01_cb10663_c1/f1p0/2458 | 3 | 1.87 | 0.50 | 0.46 | 0.40 |
Pa-eSSR-21 | F01_cb15691_c2/f1p0/2049 | 3 | 2.41 | 0.50 | 0.58 | 0.50 |
Pa-eSSR-22 | F01_cb3034_c12/f2p0/2705 | 5 | 2.85 | 0.67 | 0.65 | 0.60 |
Pa-eSSR-23 | F01_cb12182_c0/f6p2/1774 | 3 | 1.47 | 0.28 | 0.32 | 0.29 |
Pa-eSSR-24 | F01_cb13109_c0/f3p0/1635 | 5 | 2.80 | 0.48 | 0.64 | 0.58 |
Pa-eSSR-25 | F01_cb1901_c3/f1p1/2722 | 2 | 1.04 | 0.04 | 0.04 | 0.04 |
Pa-eSSR-26 | F01_cb7204_c7/f10p1/2700 | 3 | 2.65 | 0.93 | 0.62 | 0.55 |
Pa-eSSR-27 | F01_cb10594_c1/f1p0/4058 | 3 | 1.40 | 0.33 | 0.29 | 0.27 |
Pa-eSSR-28 | F01_cb9432_c36/f1p2/1811 | 5 | 1.56 | 0.43 | 0.36 | 0.33 |
Pa-eSSR-29 | F01_cb15387_c0/f3p0/1548 | 8 | 4.39 | 0.49 | 0.77 | 0.74 |
Pa-eSSR-30 | F01_cb12814_c24/f1p0/3423 | 4 | 2.67 | 0.53 | 0.62 | 0.55 |
Pa-eSSR-31 | F01_cb10835_c0/f4p0/2019 | 3 | 1.33 | 0.28 | 0.25 | 0.22 |
Total | 71 | |||||
Mean | 4.73 | 2.31 | 0.49 | 0.50 | 0.45 |
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Share and Cite
Ge, Y.; Zang, X.; Tan, L.; Wang, J.; Liu, Y.; Li, Y.; Wang, N.; Chen, D.; Zhan, R.; Ma, W. Single-Molecule Long-Read Sequencing of Avocado Generates Microsatellite Markers for Analyzing the Genetic Diversity in Avocado Germplasm. Agronomy 2019, 9, 512. https://doi.org/10.3390/agronomy9090512
Ge Y, Zang X, Tan L, Wang J, Liu Y, Li Y, Wang N, Chen D, Zhan R, Ma W. Single-Molecule Long-Read Sequencing of Avocado Generates Microsatellite Markers for Analyzing the Genetic Diversity in Avocado Germplasm. Agronomy. 2019; 9(9):512. https://doi.org/10.3390/agronomy9090512
Chicago/Turabian StyleGe, Yu, Xiaoping Zang, Lin Tan, Jiashui Wang, Yuanzheng Liu, Yanxia Li, Nan Wang, Di Chen, Rulin Zhan, and Weihong Ma. 2019. "Single-Molecule Long-Read Sequencing of Avocado Generates Microsatellite Markers for Analyzing the Genetic Diversity in Avocado Germplasm" Agronomy 9, no. 9: 512. https://doi.org/10.3390/agronomy9090512