title: IQUSP_Human_intronic_4k_v2.0

status: Public on Aug 06 2006

city: São Paulo

first: Eduardo

last: Reis

address: Av. professor Lineu Prestes, 748

department: Departamento de Bioquímica

country: Brazil

organism: Homo sapiens

description: Reporters were arranged in 12 sub-arrays containing each one 12 rows and 32 columns of spots (384 spots/sub-array). Rows 1 to 5 comprise nomatch human cDNAs. Rows 6 to 10 comprise cDNAs from known human genes. Rows 11 and 12 of each sub-array contain the 32 ScoreCard control cDNAs and 32 prostate expressed control cDNAs plus housekeeping cDNAs, respectively.

manufacturer: CAGE (Cooperation for Analysis of Gene Expression), Departamento de Bioquímica do Instituto de Química da Universidade de São Paulo.

first: Rodrigo

last: Louro

first: Helder

middle: I

last: Nakaya

first: Flavio

middle: C

last: Canavez

first: Aurea

middle: V

last: Flatschart

first: Giulliana

middle: T

last: Almeida

first: Camila

last: Egídio

first: Apuã

last: Paquola

first: Denise

last: Yamamoto

first: Renato

last: Alvarenga

first: Camille

last: da Silva

first: Aline

first: Sergio

last: Verjovski-Almeida

manufacture_protocol:
The microarray platform used in this project deploys up to 4,608 distinct cDNA spots in duplicate per glass slide, comprising a total of 9,216 elements per array. Spots represent human cDNA clones or DNA controls.
Clones were selected among the collection generated in the Human Cancer Genome Project (HCGP- http://www.ludwig.org.br/ORESTES/). A set of 1,063 ORESTES with no similarity to coding sequences of full-length human transcripts (822 Intronic, 241 New Genes). An additional set of 2,292 ORESTES clones similar to known genes were included in the microarray (Exonic), based on a list of cancer-related genes. There are 384 spots in the slide containing replicate sets of housekeeping genes (132 ), as well as negative control cDNAs (12) and sequences from characterized prostate markers genes (240). A set of 32 cDNAs from the Lucidea Microarray ScoreCard (Molecular Dynamics) was included in the microarray in each of the 12 sub-arrays (384). The remaining 485 spots are non-human contaminants cDNAs (238), empty spots (154) reserved for future use, or cDNAs that map to repeat human regions (93).
cDNA fragments deposited in the microarray are products resulting from PCR amplification of ORESTES clones, generated and sequenced in the HCGP, using universal primers. The clone collection is stored frozen in bar-coded 96-well plates, and selected clones were re-arrayed with an automated robotic operation. PCR products were purified by filtration on Multiscreen plates (Millipore cat. # MAFB NOB 50) and an aliquot was used to determine the size of the product and to quantify the amount of cDNA recovered. This was done by agarose gel electrophoresis stained with ethydium bromide and scanned on the ImageMaster Video Digital System (Amersham Pharmacia).  cDNA samples were diluted 1:1 in DMSO and spotted at high density into silane-coated, reflective Type-7 glass slides (Molecular Dynamics) using a GenIII microarray spotter robot (Molecular Dynamics). Following spotting, cDNAs were fixed to slide surface by UV cross-linking (500 mJ) and kept on a low humidity environment until use.
In this update, all spots were re-annotated according human genome hg19 assembly. Well-annotated RefSeq and UCSC genes were used and resulted in a set of 2,371 exonic coding spots, 74 known non-coding RNA spots, 188 intergenic non-coding spots, and 722 intronic non-coding spots.