Skip to main content
. 2002 Sep;12(9):1434–1444. doi: 10.1101/gr.130502

Figure 4.

Figure 4

Maintenance and amplification of pBAC/oriV that carries DNA inserts of various length. (A) Comparison of amplification of pBAC/oriV vector, with or without a 20-kb insert. After 5 h of growth, cells from the 4.5-mL volume of the culture were collected, and DNA was phenol-extracted, precipitated with 70% ethanol, digested with NcoI (lanes 1–3) or SalI (lanes 4–6), and run on an 0.8% agarose gel. (Lanes 1–3) Strain JW371 carrying pBAC/oriV grown in the LB medium (LB), LB + 0.2% D-glucose (G) or LB + 0.01% L-arabinose (A), respectively (two bands are analogous to those in Fig. 3); (lanes 4–6) strain JW378 carrying pBAC/oriV with the 20-kb insert grown, respectively, in LB, LB + 0.2% G or LB + 0.01% A. (B) Assessment of amplification by diluting of the amplified DNA of pBAC/oriV clones containing 40-kb or 80-kb inserts. Growth conditions, DNA analysis after SalI digestion, and abbreviations are as described for A and in Methods. Numbers below the lanes indicate the fold of DNA dilution prior to SalI digestion (results were similar for dilutions made after digestions and are not shown here). (Lane 1) Uninduced strain JW389 carrying pBAC/oriV with the 40-kb insert grown in LB + G; (lanes 2–5) induced strain JW389 grown in LB + A; (lane 6) uninduced strain JW390 carrying pBAC/oriV with the 80-kb insert grown in LB + G; (lanes 7–10) induced strain JW390 grown in LB + A. The DNA in lanes 1, 2, 6, and 7 is undiluted. In lanes 3–5 and 8–10, the DNA was diluted, as specified below the lanes.