Sample preparation for alfalfa relative DNA content analysis by flow cytometry

In order to select optimum extract methods for high purity alfalfa nuclear suspensions for flow cytometry(FCM), the nuclei suspension of Huaiyin alfalfa (Medicago sativa cv. huaiyin) was isolated either mechanically or by mixed enzyme solutions. The appropriate preparation method was selected based on the suspensions quality. LB01 buffer can extract larger number of nuclei with intact cell morphology and internal structure which performed best among these 5 buffers. Both of the nuclear suspensions prepared by two mixed enzyme solutions showed clear peaks on the histograms and had more nucleus. The nuclear suspension prepared by enzyme solution I was better than enzyme solution Ⅱ. However, the enzymatic methods were time-consuming and produced more impurities in the nuclear suspension. In conclusion, the mechanically extraction with LB01 buffer was appropriate method to prepare nulear suspensions for alfalfa nuclear DNA content analysis by FCM.