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. 2012 Aug 30;40(20):10375–10383. doi: 10.1093/nar/gks823

Figure 3.

Figure 3.

CHEF gel analysis of a Prochlorococcus MED4 whole genome (clone 1–13) in W303-1 A. Agarose plugs were prepared from wild-type Prochlorococcus MED4 cells (Pro), wild-type W303-1 A yeast cells (Y) and W303-1 A Prochlorococcus MED4 clone 1–13 cells and treated to isolate genomic DNA. Plugs containing only wt Prochlorococcus genomic DNA were digested with AsiSI for whole linear genomes (Pro, lane 2) or RsrII for half-genome fragments (Pro, lane 6). Plugs containing wt W303-1 A yeast cells or clone 1–13 yeast cells were digested with AscI, FseI, MreI and SfiI. These enzymes cleave yeast chromosomes but do not have recognition sites in the Prochlorococcus MED4 genome. Electrophoresis of plugs at constant voltage (100 V) for 90 min in a 1% Tris Acetate EDTA (TAE) gel removed linear yeast chromosomal DNA, while large circular Prochlorococcus DNA molecules remain topologically trapped in the plugs. Clone 1–13 or wt W303-1 A plugs were then digested with the single-cutter NotI (only present in the cloning vector of clone 1–13, lane 4) or RsrII (1–13, lane 8). RsrII cuts the Prochlorococcus MED4 genome twice into 827 and 830 kb fragments. Yeast chromosome marker (M).