Abstract:

The amplification between two adjacent MITEs was called Inter-MITE polymorphism (IMP). To meet the demand of tobacco genetic analysis and establishment of stable IMP amplification method, the IMP PCR system was optimized using IRD primer. In this study, every major amplification parameter such as template, Mg2+, dNTP, rTaq concentration were optimized analysis by Li-cor 4300 system based on tobacco genome DNA. Result showed the optimized PCR-amplifications were performed in a 20 μL volume containing 70 ng template DNA, 2.0 mmol/L of MgCl2+, 0.2 mmol/L of dNTPs, 1.0 μmol/L of labeled primer and 1.5 units of rTaq DNA polymerase. The result demonstrates that the optimized IMP method is stable robust and can be effectively applied in Tobacco genetic diversity analysis.