2.4. DNA templates used in capture experiments
For the LASSO probe capture experiments sensitivity test and positive
controls for capture, we used a 7,249 bp ssDNA isolated from the M13mp18
phage (NEB). For the capture experiments of the E. coli ORFeome by LASSO
probes, total genomic DNA of the E. coli strain K12, substrain W3110
(Migula) Castellani and Chalmers (ATCC 27325) was extracted from 500 μl
of LB broth (Sigma Aldrich) overnight culture using a Charge Switch
genomic DNA Mini Bacteria Kit (Life Technology). Sheared total genomic
DNA of E. coli K12 was obtained by sonicating 1 μg of total DNA in a
volume of 200 μl in a 1.5 ml Eppendorf tube on ice using a Branson
sonifier 450 (VWR Scientific) at an output control of 2 and a duty cycle
of 50% for 40 s.