In-House Microarrays

In-House Microarrays

 

Step 1

Identification of the genes to be arrayed, based on specific needs. (Efficient use of the array space)

Step 2

The entire gene data collection is maintained as plasmids in bacterial colonies.

Plasmid amplification (PCR to amplify the DNA in the plasmid insert). Each PCR product corresponds to a gene segment to be placed on the microarray (PCR quality control)

Step 3

The PCR-amplified DNA is arrayed onto glass slides or nylon membranes. It is necessary to test the amount of DNA, using DNA-binding dyes, for proving the uniform distribution of DNA across the array...

Step 4

Probing:

  • extraction of total RNA or m RNA
  • c-DNA
  • cDNA labeled with 33P for use with nylon membrane arrays or cDNA labeled with fluorescent dyes for glass slide arrays
  • checking the level of incorporation  of dye or radioactivity into the cDNA
  • washing off the unhybridized DNA
  • The nylon or glass arrays are scanned for radioactivity or fluorescence
  • the resulting image files are submitted to a bioinformatics process far data analysis.

References:

 "In-House Microarrays Put Researchers in Control" by J. Boguslavsky, in : Drug Discovery & Development, October 2000, 30 - 36

"Do-it-Yourself Chips" at http://www.febit.com

"Renewable Surface Biosensors" by Cynthia J. Bruckner-Lea, Darrell P. Chandler, Jay W. Grate at ISUP 2002 - HTS & POC (International Symposium on Ultramicrochemical Processes 2002 - High Throughput Screening & Process-on-chip) - Seoul, Korea, February 25 - 26 (2002) ( http://cups.kaist.ac.kr/cups/isup.html )