Licorice Germplasm Resources Identification Using DNA Barcodes Inner-Variants
Abstract
:1. Introduction
2. Results
2.1. Morphological Characteristics of Licorice Seeds
2.2. Sanger Sequencing of DNA Barcoding
2.3. Full-Length Amplicon Sequencing of ITS2, psbA-trnH, and ITS
3. Discussion
4. Materials and Methods
4.1. Sample Collection of Licorice Seeds
4.2. Total DNA Extraction, PCR Amplification and Sanger Sequencing
4.3. NGS and SMRT Sequencing
4.4. Sequencing Results and Data Analysis
5. Conclusions
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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1. ITS2 | variable site/bp | ||||
haplotype | 16 | 17 | 18 | Species | Sample |
I2-i | T | G | C | G. uralensis | GGY1, GGY2, GGE1, GGE2, GGE3, WGC, CFG, XJG |
I2-ii | C | A | A | G. glabra/inflata | GuaG, ZhaG |
2. ITS | variable site/bp | ||||
haplotype | 187 | 411~413 | Species | Sample | |
I-i | C | TGC | G. uralensis | GGY1, GGY2, GGE1, GGE2, GGE3, WGC, CFG, XJG | |
I-ii | T | CAA | G. glabra/inflata | GuaG, ZhaG | |
3. psbA-trnH | variable site/bp | ||||
haplotype | 189 | 235 | 288 | Species | Sample |
PT-i | A | C | G | G. uralensis | GGY1, GGE1, GGE2, CFG |
PT-ii | A | T | G | G. uralensis | GGY2, GGE3,WGC, XJG |
PT-iii | C | T | G | G. glabra | GuaG |
PT-iv | A | T | A | G. inflata | ZhaG |
Total Reads | Raw Data | Merged | Filtered | Mean Length/bp |
---|---|---|---|---|
ITS2 | 1,387,176 | 1,246,275 | 1,199,501 | 407 |
psbA-trnH | 1,128,832 | 1,120,013 | 1,082,034 | 389 |
Sub Reads | Circular Consensus Sequence Filtration | Mean Length/bp | ||
ITS | 14,540,927 | 218,721 | 736 |
Sample | Organ | Specie | Wild/Cultivated | Description |
---|---|---|---|---|
GGY1 | Seeds | G. uralensis | Cultivated | batch 1 of Market A |
GGY2 | Seeds | G. uralensis | Cultivated | batch 2 of Market A |
GGE1 | Seeds | G. uralensis | Cultivated | batch 1 of Market B |
GGE2 | Seeds | G. uralensis | Cultivated | batch 2 of Market B |
GGE3 | Seeds | G. uralensis | Cultivated | batch 2 of Market B |
GuaG | Seeds | G. glabra | Wild | Market C |
ZhaG | Seeds | G. inflata | Wild | Market C |
WGC | Seeds | G. uralensis | Wild | Market C |
CFG | Seeds | G. uralensis | Wild | Chifeng, Inner Mongolia, China |
XJG | Seeds | G. uralensis | Wild | Xinjiang, China |
Region | Primer Sequence | Program |
---|---|---|
ITS2 | ITS2-P3: YGACTCTCGGCAACGGATA ITS2-E4: RGTTTCTTTTCCTCCGCTTA [44] | 94 °C for 5 min; 94 °C for 1 min, 55 °C for 1 min, 72 °C for 1.5 min, 30 cycles; and 72 °C for 7 min |
psbA-trnH | psbAF: GTTATGCATGAACGTAATGCTC trnHR: CGCGCATGGTGGATTCACAATCC [16] | 94 °C for 5 min; 94 °C for 1 min, 55 °C for 1 min, 72 °C for 1.5 min, 30 cycles; and 72 °C for 7 min |
ITS | ITS4:TCCTCCGCTTATTGATATGC ITS5:GGAAGTAAAAGTCGTAACAAGG [58] | 95 °C for 3 min; 95 °C for 30 s, 55 °C for 30 s, 72 °C for 1.5 min, 35 cycles; and 72 °C for 10 min |
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Liu, Q.; Guo, S.; Zheng, X.; Shen, X.; Zhang, T.; Liao, B.; He, W.; Hu, H.; Cheng, R.; Xu, J. Licorice Germplasm Resources Identification Using DNA Barcodes Inner-Variants. Plants 2021, 10, 2036. https://doi.org/10.3390/plants10102036
Liu Q, Guo S, Zheng X, Shen X, Zhang T, Liao B, He W, Hu H, Cheng R, Xu J. Licorice Germplasm Resources Identification Using DNA Barcodes Inner-Variants. Plants. 2021; 10(10):2036. https://doi.org/10.3390/plants10102036
Chicago/Turabian StyleLiu, Qianwen, Shuai Guo, Xiasheng Zheng, Xiaofeng Shen, Tianyi Zhang, Baosheng Liao, Wenrui He, Haoyu Hu, Ruiyang Cheng, and Jiang Xu. 2021. "Licorice Germplasm Resources Identification Using DNA Barcodes Inner-Variants" Plants 10, no. 10: 2036. https://doi.org/10.3390/plants10102036
APA StyleLiu, Q., Guo, S., Zheng, X., Shen, X., Zhang, T., Liao, B., He, W., Hu, H., Cheng, R., & Xu, J. (2021). Licorice Germplasm Resources Identification Using DNA Barcodes Inner-Variants. Plants, 10(10), 2036. https://doi.org/10.3390/plants10102036