title: CAB-MEL-LfeRT32a-14000-V2
status: Public on Jun 23 2016
contact.city: Torrejón de Ardoz
contact.first: Mercedes
contact.last: Moreno
contact.state: Madrid
contact.address: Crta. de Ajalvir, km 4
contact.country: Spain
contact.institute: Centro de Astrobiología
organism: Leptospirillum ferrooxidans
manufacture_protocol: 1. A random genomic library from Leptospirillum ferrooxidans RT32a was created as described in Parro V. and Moreno-Paz M. (PNAS, 100(13):7883-7888).
  2. Clone fragments were amplified by PCR and purified with a PCR Microarray Purification Kit for 96-Well and 50 µl PCR samples (Arrayit Corporation).
  3. Purified PCR products were dried in a Universal Vacuum System Plus UVS400A (Savant) and resuspended in 15ul of water. After resuspension, 15ul of 2x Spotting Solution Plus (Arrayit Corporation) was added into 384-V-bottom plates from Genetix (cat# X6004).
  4. 15ul aliquots were dispensed into 384-V-bottom plates from Genetix (cat# X6004) with the Epmotion 5075vac system (Eppendorf)
  5. Plates were shaked near maximum rotational speed on flat-bed shaker for 5 minutes.
  6. Plates were centrifugated at 2000 rpm for 3 minutes.
  7. Plates were placed in the appropriate location of plate cassette before printing run.
  8. After run, plates were sealed with cover plates and maintained in -20ºC.