By R. Allen Dyer
This article is a pictorial presentation and contains a key to genera and species.
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The fourth round stacking was perfumed as for the second round stacking. Nine out of 20 chloramphenicol and ampicillin colonies show the PCR product when amplified by primer sets p + h and g + f, consistent with the molecule ABCDE structure (Fig. 5a, b). 8 kb bands (Fig. 5c). Prior to transformation into E. coli, representative molecule ABCDE was incubated with Cre recombiase cell extracts. Three out of 36 colonies were chloramphenicol resistant but ampicillin sensitive. 4 kb deletion junction amplified by primer sets x + z which is consistent with ABCDE− structure (Fig.
7. 5 mg/mL X-Gluc. 2 Tobacco Medium Composition 1. Basic medium (MS0): Add 100 mL of 10× MS macro salts, 10 mL of 100× MS micro salts and vitamins, 10 mL of 100× FeEDTA, 30 g/L sucrose to 800 mL distilled water, and bring volume up to 1 L. 8 and add 4 g/L phytagel. 2. 1 mL of 1 mg/mL NAA (add after autoclaving). 3. Selection medium (MS2): MS1 plus 2 mL of 40 mg/mL kanamycin (add after autoclaving). 24 Ruyu Li et al. 5 Materials for Bombardment 1. PDS-1000/He particle delivery system from Bio-Rad (Hercules, CA, USA).
2 μL retrieved DNA is transformed into 50 μL E. coli DH5α competent cells and selected on chloramphenicol and ampicillin supplemented LB plates. 5. 20 single clones is selected, each into 10 μL sterilized ddH2O for 30 cycles of colony PCR in 20 μL reaction volume containing 10 μL 2× Taq Master Mix, 1 μL colony template, 10 μM each primers and sterile ddH2O. 6. PCR product is separated on 1 % agarose gel. Of the 20 colonies resistant to chloramphenicol and ampicillin, four clones (20 %) showed PCR products amplified by primer sets b + d and a + c consistent with molecule AB structure, two clones showed PCR products amplified by primer b + d2 and a2 + c, consistent with structure AB′ (Fig.