Fig. 4

Overview of isothermal DNA construction on the microfluidics platform. a Overview of the basic IHDC step on the microfluidics platform. Stage I. Two oligos A and B (as shown, or alternatively two DNA fragments A and B) and a mixture of enzymes are transferred to the reactor. Stage II. Primers P1 and P2, and a mixture of enzymes, are transferred to the reactor. Stage III. A mixture of ATP and magnesium acetate, and a mixture of enzymes, are transferred to the reactor. The temperature is increased to 38 °C, and the reaction is incubated for 15 min. As a result, an elongated and amplified DNA fragment AB is produced. b Hierarchical construction tree of seven separate synthesis reactions that result in the final product (gfp as shown). c Gel electrophoresis image of all the intermediates and the final gfp construct. Lanes as labelled by: M: GeneRuler 1 kb Plus DNA Ladder (Thermo Scientific); Level 1 (quarter) fragments: 1-2, 3-4, 5-6, 7-8; Level 2 (half) fragments: 1-4 and 5-8; Level 3 (full length gfp) fragment: 1-8