title: AgResearch Bos taurus Clone Array Print 114
status: Public on Feb 20 2008
contact.city: Puddle Alley
contact.first: Alan
contact.middle: Francis
contact.last: McCulloch
contact.email: alan.mcculloch@agresearch.co.nz
contact.state: Mosgiel
contact.address: Invermay
contact.country: New Zealand
contact.institute: AgResearch NZ
contact.phone: 64 3 4899080
organism: Bos taurus
manufacture_protocol: Microarray preparation
  The Bovine ESTs used for the cDNA microarry  were made and sequenced by the Genesis Company Auckland NZ . The 23K array consisted  of  EST  from 50 tissue libraries covering  brain, mammary, heart and skeletal muscle, glands, ovaries, fat, liver, intestine, spleen and PBM cells..
  Clones in pBK-CMV vector were amplified by 55pmol primers T7 anad T3 and those in pcMV-sport6 by 45pmol T7 and Sp6  in a 50ul reaction with 0.2mM dNTPs, 1.5mM mgCl and 2.5 units AB red hot taq polymerase in 96 well plates.  The  PCR was carried out at 55ºC for 36 cycles and extended at 72ºC for 5min.
  The product was verified by running a portion on a 1% gel. The sample were then purified by isopropanol and ETOH precipitation.
  
  The cDNA samples were suspended in 5ul 3x SSC giving a concentration of 100 to 200ng.
  Printing was performed on poly L Lysine coated slides using split pin technology on the ESI array robot with a 32 pin configuration.