Development and application of perfect SSR markers in cotton
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Journal of Cotton Research
RESEARCH
Open Access
Development and application of perfect SSR markers in cotton WU Yuzhen, HUANG Longyu, ZHOU Dayun, FU Xiaoqiong, LI Chao, WEI Shoujun, PENG Jun* and KUANG Meng*
Abstract Background: This study aimed to develop a set of perfect simple sequence repeat (SSR) markers with a single copy in the cotton genome, to construct a DNA fingerprint database suitable for authentication of cotton cultivars. We optimized the polymerase chain reaction (PCR) system for multi-platform compatibility and improving detection efficiency. Based on the reference genome of upland cotton and 10× resequencing data of 48 basic cotton germplasm lines, single-copy polymorphic SSR sites were identified and developed as diploidization SSR markers. The SSR markers were detected by denaturing polyacrylamide gel electrophoresis (PAGE) for initial screening, then fluorescence capillary electrophoresis for secondary screening. The final perfect SSR markers were evaluated and verified using 210 lines from different sources among Chinese cotton regional trials. Results: Using bioinformatics techniques, 1 246 SSR markers were designed from 26 626 single-copy SSR loci. Adopting a stepwise (primary and secondary) screening strategy, a set of 60 perfect SSR markers was selected with high amplification efficiency and stability, easy interpretation of peak type, multiple allelic variations, high polymorphism information content (PIC) value, uniform chromosome distribution, and single-copy characteristics. A multiplex PCR system was established with ten SSR markers using capillary electrophoresis detection. Conclusions: A set of perfect SSR markers of cotton was developed and a high-throughput SSR marker detection system was established. This study lays a foundation for large-scale and standardized construction of a cotton DNA fingerprint database for authentication of cotton varieties. Keywords: Cotton, SSR marker, Cultivar identification, DNA fingerprint database
Background Cotton is one of the most important economic crops in the world. Its fiber is a renewable textile raw material, and its seeds can be used for the production and processing of cottonseed oil. In recent years, excellent new varieties have been developed and promoted, and the seed market has flourished. At the same time, brand infringement and the production and sale of fake and inferior seeds have continued to occur, and this has severely dampened the enthusiasm of innovators, increased the risks for farmers, and affected agricultural * Correspondence: [email protected]; [email protected] Institute of Cotton Research, Chinese Academy of Agricultural Sciences / Zhengzhou Research Base, State Key Laboratory of Cotton Biology, Zhengzhou University, Zhengzhou 455001, China
production safety, which has severely restricted the development of China’s seed industry (Wu et al. 2015). Variety identification can effectively ensure the quality of seeds in agricultural production and reduce the risk of using fake and/or inferior seeds. Among DNA molecu
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