Articles
THE ORTHOGONAL OPTIMIZATION OF SRAP AMPLIFICATION SYSTEM IN CHINESE JUJUBE
Article number
840_16
Pages
143 – 148
Language
English
Abstract
Sequence-related amplified polymorphism (SRAP) is a new marker technique with the characteristics of simplicity, reliability, moderate through-put ratio and facile sequencing of selected bands.
The orthogonal design was used to optimize SRAP-PCR amplification system on Chinese jujube in three levels of four factors (dNTP, primer, Mg2+, Taq DNA polymerase, and DNA template) respectively.
The results showed that the orthogonal experiment design was a practicable method and in a total volume of 20 µl SRAP-PCR system, it contains 1×buffer, 200 µmol/L dNTP, 30 ng Primer, 2.5 mmol/L MgCl2, 1.0 µ Taq DNA polymerase and 20 ng template DNA. The suitable PCR procedure was initially denaturing at 94°C for 5 min; then 94°C for 1 min, 33°C for l min, and 72°C for 1 min for the first five cycles; then the annealing temperature is raised to 53°C for another 30 cycles; 72°C for 5 min for the final extension.
The new established SRAP-PCR amplification system and procedure of Chinese jujube had good repeatability and stability, and also satisfied the genome amplification requests of Ziziphus. SRAP-PCR system could be applied in genetic relationships between various groups in Ziziphus.
The orthogonal design was used to optimize SRAP-PCR amplification system on Chinese jujube in three levels of four factors (dNTP, primer, Mg2+, Taq DNA polymerase, and DNA template) respectively.
The results showed that the orthogonal experiment design was a practicable method and in a total volume of 20 µl SRAP-PCR system, it contains 1×buffer, 200 µmol/L dNTP, 30 ng Primer, 2.5 mmol/L MgCl2, 1.0 µ Taq DNA polymerase and 20 ng template DNA. The suitable PCR procedure was initially denaturing at 94°C for 5 min; then 94°C for 1 min, 33°C for l min, and 72°C for 1 min for the first five cycles; then the annealing temperature is raised to 53°C for another 30 cycles; 72°C for 5 min for the final extension.
The new established SRAP-PCR amplification system and procedure of Chinese jujube had good repeatability and stability, and also satisfied the genome amplification requests of Ziziphus. SRAP-PCR system could be applied in genetic relationships between various groups in Ziziphus.
Publication
Authors
L. Li, J.Y. Peng, R.H. Huang, R.X. Bai, B.Q. Zheng
Keywords
Chinese jujube, SRAP, system optimization, orthogonal
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