Isolation of chromosome DNA of wheat blue dwarf phytoplasma

Abstract:

[Objective] Isolating chromosome DNA of wheat blue dwarf (WBD) phytoplasma and establishing an effective protocol of purification for chromosome DNA. [Methods] Dif-ferential centrifugation and plus-filed gel electrophoresis (PFGE) were used to purify the chromosome DNA of WBD phytoplasma. The band observed in PFGE was confirmed by PCR and Southern blot. The effect of each step was detected by real-time quantitative PCR analysis. [Results] Using differential centrifugation and PFGE, a band about 650 kb was observed, which was confirmed as the chromosome DNA of WBD phytoplasma by Southern blot hy-bridization and PCR. Meanwhile, real-time PCR analysis showed that the relative copies of WBD phytoplasma chromosome DNA derived from differential centrifugation and PFGE was 436.5 times than that in the total DNA of infected periwinkle. [Conclusion] The size of WBD phytoplasma chromosome DNA is about 650 kb, purified WBD phytoplasma chromosome DNA can be obtained effectively using differential centrifugation and PFGE.