Detail information
ID ENOM00071
Accession GSE9525
Status Public on Oct 01, 2008
Title Effect of short- and long-term morphine treatment on gene expression in the hypothalamus and pituitary
Organism Mus musculus
Experiment type Expression profiling by array
Summary
Addictive drugs including opioids activate signal transduction pathways that regulate gene expression in the brain. However, changes in CNS gene expression following morphine exposure are poorly understood. We studied the effect of short- and long-term morphine treatment on gene expression in the hypothalamus and pituitary using genome-wide DNA microarray and real-time reverse transcriptase polymerase chain reaction (RT-PCR) analyses. In the hypothalamus, we found that short-term morphine administration up-regulated (at least 2-fold) 39 genes and down-regulated six genes. Long-term morphine administration up-regulated 35 genes and down-regulated 51 hypothalamic genes. In the pituitary, we found that short-term morphine administration up-regulated (at least 2-fold) 110 genes and down-regulated 29 genes. Long-term morphine administration up-regulated 85 genes and down-regulated 37 pituitary genes. Strikingly, microarray analysis uncovered several genes involved in food intake (neuropeptide Y, agouti-related protein, and cocaine and amphetamine-regulated transcript) whose expression was strongly altered by morphine exposure in either the hypothalamus or pituitary. Subsequent RT-PCR analysis confirmed similar gene regulation of noteworthy genes in these regions. Finally, we found functional correlation between morphine-induced alterations in food intake and regulations of genes involved in this process. Changes in genes related to food intake may uncover new pathways related to some of the physiological effects of opioids.
Samples
GSM ID Sample info Characteristics Description
GSM239721 Hypothalamus 4day(H1/L1) Strain: C57BL/6, Gender: male, Tissue: hypothalamus;Strain: C57BL/6, Gender: male, Tissue: hypothalamus DNA microarrays were made at the SCCPRR Gene Array Facility (Department of Pharmacology, University of Washington) using the Programs for Genomic Applications (PGA) mouse 70mer oligo library generated at Massachusetts General Hospital (Boston, MA). The mRNAs to be compared are labeled with two different fluorescent cyanine dyes (Cy 3 or Cy5) using an indirect labeling method. Amino allyl-dUTP is incorporated during reverse transcription and then post-coupled with a NHS-esther mono-reactive cyanine dye. This indirect labeling method allows us to avoid one common form of false positive resulting from the fact that the reverse transcriptase enzyme differentially utilizes the Cy5 and Cy3 dyes. The two labeled cDNA samples are then combined and hybridized on a single microarray slide overnight using a standard formamide-based RNA hybridization protocol (see Protocols toggle on website). The following morning the slide is washed, dried, and scanned using the Axon 4000a scanner. The GenePix® software on the Axon 4000a scanner, after simultaneous laser scanning dual excitation/emission capturing the Cy5 and Cy3 channels, creates a merged 16-bit TIFF image, finds the spots, performs a background subtraction/signal to noise analysis, and then generates a .gpr file containing a variety of parameters concerning each individual spot.
GSM241212 Hypothalamus 4day(H1s/L1s);dye swap Strain: C57BL/6, Gender: male, Tissue: hypothalamus;Strain: C57BL/6, Gender: male, Tissue: hypothalamus DNA microarrays were made at the SCCPRR Gene Array Facility (Department of Pharmacology, University of Washington) using the Programs for Genomic Applications (PGA) mouse 70mer oligo library generated at Massachusetts General Hospital (Boston, MA). The mRNAs to be compared are labeled with two different fluorescent cyanine dyes (Cy 3 or Cy5) using an indirect labeling method. Amino allyl-dUTP is incorporated during reverse transcription and then post-coupled with a NHS-esther mono-reactive cyanine dye. This indirect labeling method allows us to avoid one common form of false positive resulting from the fact that the reverse transcriptase enzyme differentially utilizes the Cy5 and Cy3 dyes. The two labeled cDNA samples are then combined and hybridized on a single microarray slide overnight using a standard formamide-based RNA hybridization protocol (see Protocols toggle on website). The following morning the slide is washed, dried, and scanned using the Axon 4000a scanner. The GenePix® software on the Axon 4000a scanner, after simultaneous laser scanning dual excitation/emission capturing the Cy5 and Cy3 channels, creates a merged 16-bit TIFF image, finds the spots, performs a background subtraction/signal to noise analysis, and then generates a .gpr file containing a variety of parameters concerning each individual spot.
GSM241213 Hypothalamus 4day(H2/L2);biological replicate Strain: C57BL/6, Gender: male, Tissue: hypothalamus;Strain: C57BL/6, Gender: male, Tissue: hypothalamus DNA microarrays were made at the SCCPRR Gene Array Facility (Department of Pharmacology, University of Washington) using the Programs for Genomic Applications (PGA) mouse 70mer oligo library generated at Massachusetts General Hospital (Boston, MA). The mRNAs to be compared are labeled with two different fluorescent cyanine dyes (Cy 3 or Cy5) using an indirect labeling method. Amino allyl-dUTP is incorporated during reverse transcription and then post-coupled with a NHS-esther mono-reactive cyanine dye. This indirect labeling method allows us to avoid one common form of false positive resulting from the fact that the reverse transcriptase enzyme differentially utilizes the Cy5 and Cy3 dyes. The two labeled cDNA samples are then combined and hybridized on a single microarray slide overnight using a standard formamide-based RNA hybridization protocol (see Protocols toggle on website). The following morning the slide is washed, dried, and scanned using the Axon 4000a scanner. The GenePix® software on the Axon 4000a scanner, after simultaneous laser scanning dual excitation/emission capturing the Cy5 and Cy3 channels, creates a merged 16-bit TIFF image, finds the spots, performs a background subtraction/signal to noise analysis, and then generates a .gpr file containing a variety of parameters concerning each individual spot.
GSM241214 Hypothalamus 4day(H2s/L2s);biological replicate_dye swap Strain: C57BL/6, Gender: male, Tissue: hypothalamus;Strain: C57BL/6, Gender: male, Tissue: hypothalamus DNA microarrays were made at the SCCPRR Gene Array Facility (Department of Pharmacology, University of Washington) using the Programs for Genomic Applications (PGA) mouse 70mer oligo library generated at Massachusetts General Hospital (Boston, MA). The mRNAs to be compared are labeled with two different fluorescent cyanine dyes (Cy 3 or Cy5) using an indirect labeling method. Amino allyl-dUTP is incorporated during reverse transcription and then post-coupled with a NHS-esther mono-reactive cyanine dye. This indirect labeling method allows us to avoid one common form of false positive resulting from the fact that the reverse transcriptase enzyme differentially utilizes the Cy5 and Cy3 dyes. The two labeled cDNA samples are then combined and hybridized on a single microarray slide overnight using a standard formamide-based RNA hybridization protocol (see Protocols toggle on website). The following morning the slide is washed, dried, and scanned using the Axon 4000a scanner. The GenePix® software on the Axon 4000a scanner, after simultaneous laser scanning dual excitation/emission capturing the Cy5 and Cy3 channels, creates a merged 16-bit TIFF image, finds the spots, performs a background subtraction/signal to noise analysis, and then generates a .gpr file containing a variety of parameters concerning each individual spot.
GSM241215 Pituitary 4day(P1) Strain: C57BL/6, Gender: male, Tissue: pituitary;Strain: C57BL/6, Gender: male, Tissue: pituitary DNA microarrays were made at the SCCPRR Gene Array Facility (Department of Pharmacology, University of Washington) using the Programs for Genomic Applications (PGA) mouse 70mer oligo library generated at Massachusetts General Hospital (Boston, MA). The mRNAs to be compared are labeled with two different fluorescent cyanine dyes (Cy 3 or Cy5) using an indirect labeling method. Amino allyl-dUTP is incorporated during reverse transcription and then post-coupled with a NHS-esther mono-reactive cyanine dye. This indirect labeling method allows us to avoid one common form of false positive resulting from the fact that the reverse transcriptase enzyme differentially utilizes the Cy5 and Cy3 dyes. The two labeled cDNA samples are then combined and hybridized on a single microarray slide overnight using a standard formamide-based RNA hybridization protocol (see Protocols toggle on website). The following morning the slide is washed, dried, and scanned using the Axon 4000a scanner. The GenePix® software on the Axon 4000a scanner, after simultaneous laser scanning dual excitation/emission capturing the Cy5 and Cy3 channels, creates a merged 16-bit TIFF image, finds the spots, performs a background subtraction/signal to noise analysis, and then generates a .gpr file containing a variety of parameters concerning each individual spot.
GSM241216 Pituitary 4day(P1s);dye swap Strain: C57BL/6, Gender: male, Tissue: pituitary;Strain: C57BL/6, Gender: male, Tissue: pituitary DNA microarrays were made at the SCCPRR Gene Array Facility (Department of Pharmacology, University of Washington) using the Programs for Genomic Applications (PGA) mouse 70mer oligo library generated at Massachusetts General Hospital (Boston, MA). The mRNAs to be compared are labeled with two different fluorescent cyanine dyes (Cy 3 or Cy5) using an indirect labeling method. Amino allyl-dUTP is incorporated during reverse transcription and then post-coupled with a NHS-esther mono-reactive cyanine dye. This indirect labeling method allows us to avoid one common form of false positive resulting from the fact that the reverse transcriptase enzyme differentially utilizes the Cy5 and Cy3 dyes. The two labeled cDNA samples are then combined and hybridized on a single microarray slide overnight using a standard formamide-based RNA hybridization protocol (see Protocols toggle on website). The following morning the slide is washed, dried, and scanned using the Axon 4000a scanner. The GenePix® software on the Axon 4000a scanner, after simultaneous laser scanning dual excitation/emission capturing the Cy5 and Cy3 channels, creates a merged 16-bit TIFF image, finds the spots, performs a background subtraction/signal to noise analysis, and then generates a .gpr file containing a variety of parameters concerning each individual spot.
GSM241217 Pituitary 4day(P2/L1);biological replicate Strain: C57BL/6, Gender: male, Tissue: pituitary;Strain: C57BL/6, Gender: male, Tissue: pituitary DNA microarrays were made at the SCCPRR Gene Array Facility (Department of Pharmacology, University of Washington) using the Programs for Genomic Applications (PGA) mouse 70mer oligo library generated at Massachusetts General Hospital (Boston, MA). The mRNAs to be compared are labeled with two different fluorescent cyanine dyes (Cy 3 or Cy5) using an indirect labeling method. Amino allyl-dUTP is incorporated during reverse transcription and then post-coupled with a NHS-esther mono-reactive cyanine dye. This indirect labeling method allows us to avoid one common form of false positive resulting from the fact that the reverse transcriptase enzyme differentially utilizes the Cy5 and Cy3 dyes. The two labeled cDNA samples are then combined and hybridized on a single microarray slide overnight using a standard formamide-based RNA hybridization protocol (see Protocols toggle on website). The following morning the slide is washed, dried, and scanned using the Axon 4000a scanner. The GenePix® software on the Axon 4000a scanner, after simultaneous laser scanning dual excitation/emission capturing the Cy5 and Cy3 channels, creates a merged 16-bit TIFF image, finds the spots, performs a background subtraction/signal to noise analysis, and then generates a .gpr file containing a variety of parameters concerning each individual spot.
GSM241218 Pituitary 4day(P2s/L1s);biological replicate_dye swap Strain: C57BL/6, Gender: male, Tissue: pituitary;Strain: C57BL/6, Gender: male, Tissue: pituitary DNA microarrays were made at the SCCPRR Gene Array Facility (Department of Pharmacology, University of Washington) using the Programs for Genomic Applications (PGA) mouse 70mer oligo library generated at Massachusetts General Hospital (Boston, MA). The mRNAs to be compared are labeled with two different fluorescent cyanine dyes (Cy 3 or Cy5) using an indirect labeling method. Amino allyl-dUTP is incorporated during reverse transcription and then post-coupled with a NHS-esther mono-reactive cyanine dye. This indirect labeling method allows us to avoid one common form of false positive resulting from the fact that the reverse transcriptase enzyme differentially utilizes the Cy5 and Cy3 dyes. The two labeled cDNA samples are then combined and hybridized on a single microarray slide overnight using a standard formamide-based RNA hybridization protocol (see Protocols toggle on website). The following morning the slide is washed, dried, and scanned using the Axon 4000a scanner. The GenePix® software on the Axon 4000a scanner, after simultaneous laser scanning dual excitation/emission capturing the Cy5 and Cy3 channels, creates a merged 16-bit TIFF image, finds the spots, performs a background subtraction/signal to noise analysis, and then generates a .gpr file containing a variety of parameters concerning each individual spot.
GSM241569 Hypothalamus 6hour(H1/S1) Strain: C57BL/6, Gender: male, Tissue: hypothalamus;Strain: C57BL/6, Gender: male, Tissue: hypothalamus DNA microarrays were made at the SCCPRR Gene Array Facility (Department of Pharmacology, University of Washington) using the Programs for Genomic Applications (PGA) mouse 70mer oligo library generated at Massachusetts General Hospital (Boston, MA). The mRNAs to be compared are labeled with two different fluorescent cyanine dyes (Cy 3 or Cy5) using an indirect labeling method. Amino allyl-dUTP is incorporated during reverse transcription and then post-coupled with a NHS-esther mono-reactive cyanine dye. This indirect labeling method allows us to avoid one common form of false positive resulting from the fact that the reverse transcriptase enzyme differentially utilizes the Cy5 and Cy3 dyes. The two labeled cDNA samples are then combined and hybridized on a single microarray slide overnight using a standard formamide-based RNA hybridization protocol (see Protocols toggle on website). The following morning the slide is washed, dried, and scanned using the Axon 4000a scanner. The GenePix® software on the Axon 4000a scanner, after simultaneous laser scanning dual excitation/emission capturing the Cy5 and Cy3 channels, creates a merged 16-bit TIFF image, finds the spots, performs a background subtraction/signal to noise analysis, and then generates a .gpr file containing a variety of parameters concerning each individual spot.
GSM241570 Hypothalamus 6hour(H1s/S1s);dye swap Strain: C57BL/6, Gender: male, Tissue: hypothalamus;Strain: C57BL/6, Gender: male, Tissue: hypothalamus DNA microarrays were made at the SCCPRR Gene Array Facility (Department of Pharmacology, University of Washington) using the Programs for Genomic Applications (PGA) mouse 70mer oligo library generated at Massachusetts General Hospital (Boston, MA). The mRNAs to be compared are labeled with two different fluorescent cyanine dyes (Cy 3 or Cy5) using an indirect labeling method. Amino allyl-dUTP is incorporated during reverse transcription and then post-coupled with a NHS-esther mono-reactive cyanine dye. This indirect labeling method allows us to avoid one common form of false positive resulting from the fact that the reverse transcriptase enzyme differentially utilizes the Cy5 and Cy3 dyes. The two labeled cDNA samples are then combined and hybridized on a single microarray slide overnight using a standard formamide-based RNA hybridization protocol (see Protocols toggle on website). The following morning the slide is washed, dried, and scanned using the Axon 4000a scanner. The GenePix® software on the Axon 4000a scanner, after simultaneous laser scanning dual excitation/emission capturing the Cy5 and Cy3 channels, creates a merged 16-bit TIFF image, finds the spots, performs a background subtraction/signal to noise analysis, and then generates a .gpr file containing a variety of parameters concerning each individual spot.
GSM241571 Hypothalamus 6hour (H2/S2);biological replicate Strain: C57BL/6, Gender: male, Tissue: hypothalamus;Strain: C57BL/6, Gender: male, Tissue: hypothalamus DNA microarrays were made at the SCCPRR Gene Array Facility (Department of Pharmacology, University of Washington) using the Programs for Genomic Applications (PGA) mouse 70mer oligo library generated at Massachusetts General Hospital (Boston, MA). The mRNAs to be compared are labeled with two different fluorescent cyanine dyes (Cy 3 or Cy5) using an indirect labeling method. Amino allyl-dUTP is incorporated during reverse transcription and then post-coupled with a NHS-esther mono-reactive cyanine dye. This indirect labeling method allows us to avoid one common form of false positive resulting from the fact that the reverse transcriptase enzyme differentially utilizes the Cy5 and Cy3 dyes. The two labeled cDNA samples are then combined and hybridized on a single microarray slide overnight using a standard formamide-based RNA hybridization protocol (see Protocols toggle on website). The following morning the slide is washed, dried, and scanned using the Axon 4000a scanner. The GenePix® software on the Axon 4000a scanner, after simultaneous laser scanning dual excitation/emission capturing the Cy5 and Cy3 channels, creates a merged 16-bit TIFF image, finds the spots, performs a background subtraction/signal to noise analysis, and then generates a .gpr file containing a variety of parameters concerning each individual spot.
GSM241572 Hypothalamus 6hour (H2s/S2s);biological replicate_dye swap Strain: C57BL/6, Gender: male, Tissue: hypothalamus;Strain: C57BL/6, Gender: male, Tissue: hypothalamus DNA microarrays were made at the SCCPRR Gene Array Facility (Department of Pharmacology, University of Washington) using the Programs for Genomic Applications (PGA) mouse 70mer oligo library generated at Massachusetts General Hospital (Boston, MA). The mRNAs to be compared are labeled with two different fluorescent cyanine dyes (Cy 3 or Cy5) using an indirect labeling method. Amino allyl-dUTP is incorporated during reverse transcription and then post-coupled with a NHS-esther mono-reactive cyanine dye. This indirect labeling method allows us to avoid one common form of false positive resulting from the fact that the reverse transcriptase enzyme differentially utilizes the Cy5 and Cy3 dyes. The two labeled cDNA samples are then combined and hybridized on a single microarray slide overnight using a standard formamide-based RNA hybridization protocol (see Protocols toggle on website). The following morning the slide is washed, dried, and scanned using the Axon 4000a scanner. The GenePix® software on the Axon 4000a scanner, after simultaneous laser scanning dual excitation/emission capturing the Cy5 and Cy3 channels, creates a merged 16-bit TIFF image, finds the spots, performs a background subtraction/signal to noise analysis, and then generates a .gpr file containing a variety of parameters concerning each individual spot.
GSM241573 Pituitary 6hour (P1/S1) Strain: C57BL/6, Gender: male, Tissue: pituitary;Strain: C57BL/6, Gender: male, Tissue: pituitary DNA microarrays were made at the SCCPRR Gene Array Facility (Department of Pharmacology, University of Washington) using the Programs for Genomic Applications (PGA) mouse 70mer oligo library generated at Massachusetts General Hospital (Boston, MA). The mRNAs to be compared are labeled with two different fluorescent cyanine dyes (Cy 3 or Cy5) using an indirect labeling method. Amino allyl-dUTP is incorporated during reverse transcription and then post-coupled with a NHS-esther mono-reactive cyanine dye. This indirect labeling method allows us to avoid one common form of false positive resulting from the fact that the reverse transcriptase enzyme differentially utilizes the Cy5 and Cy3 dyes. The two labeled cDNA samples are then combined and hybridized on a single microarray slide overnight using a standard formamide-based RNA hybridization protocol (see Protocols toggle on website). The following morning the slide is washed, dried, and scanned using the Axon 4000a scanner. The GenePix® software on the Axon 4000a scanner, after simultaneous laser scanning dual excitation/emission capturing the Cy5 and Cy3 channels, creates a merged 16-bit TIFF image, finds the spots, performs a background subtraction/signal to noise analysis, and then generates a .gpr file containing a variety of parameters concerning each individual spot.
GSM241574 Pituitary 6hour(P1s/S1s);dye swap Strain: C57BL/6, Gender: male, Tissue: pituitary;Strain: C57BL/6, Gender: male, Tissue: pituitary DNA microarrays were made at the SCCPRR Gene Array Facility (Department of Pharmacology, University of Washington) using the Programs for Genomic Applications (PGA) mouse 70mer oligo library generated at Massachusetts General Hospital (Boston, MA). The mRNAs to be compared are labeled with two different fluorescent cyanine dyes (Cy 3 or Cy5) using an indirect labeling method. Amino allyl-dUTP is incorporated during reverse transcription and then post-coupled with a NHS-esther mono-reactive cyanine dye. This indirect labeling method allows us to avoid one common form of false positive resulting from the fact that the reverse transcriptase enzyme differentially utilizes the Cy5 and Cy3 dyes. The two labeled cDNA samples are then combined and hybridized on a single microarray slide overnight using a standard formamide-based RNA hybridization protocol (see Protocols toggle on website). The following morning the slide is washed, dried, and scanned using the Axon 4000a scanner. The GenePix® software on the Axon 4000a scanner, after simultaneous laser scanning dual excitation/emission capturing the Cy5 and Cy3 channels, creates a merged 16-bit TIFF image, finds the spots, performs a background subtraction/signal to noise analysis, and then generates a .gpr file containing a variety of parameters concerning each individual spot.
GSM241575 Pituitary 6hour(P2/S2);biological replicate Strain: C57BL/6, Gender: male, Tissue: pituitary;Strain: C57BL/6, Gender: male, Tissue: pituitary DNA microarrays were made at the SCCPRR Gene Array Facility (Department of Pharmacology, University of Washington) using the Programs for Genomic Applications (PGA) mouse 70mer oligo library generated at Massachusetts General Hospital (Boston, MA). The mRNAs to be compared are labeled with two different fluorescent cyanine dyes (Cy 3 or Cy5) using an indirect labeling method. Amino allyl-dUTP is incorporated during reverse transcription and then post-coupled with a NHS-esther mono-reactive cyanine dye. This indirect labeling method allows us to avoid one common form of false positive resulting from the fact that the reverse transcriptase enzyme differentially utilizes the Cy5 and Cy3 dyes. The two labeled cDNA samples are then combined and hybridized on a single microarray slide overnight using a standard formamide-based RNA hybridization protocol (see Protocols toggle on website). The following morning the slide is washed, dried, and scanned using the Axon 4000a scanner. The GenePix® software on the Axon 4000a scanner, after simultaneous laser scanning dual excitation/emission capturing the Cy5 and Cy3 channels, creates a merged 16-bit TIFF image, finds the spots, performs a background subtraction/signal to noise analysis, and then generates a .gpr file containing a variety of parameters concerning each individual spot.
GSM241576 Pituitary 6hour (P2s/S2s);biological replicate_dye swap Strain: C57BL/6, Gender: male, Tissue: pituitary;Strain: C57BL/6, Gender: male, Tissue: pituitary DNA microarrays were made at the SCCPRR Gene Array Facility (Department of Pharmacology, University of Washington) using the Programs for Genomic Applications (PGA) mouse 70mer oligo library generated at Massachusetts General Hospital (Boston, MA). The mRNAs to be compared are labeled with two different fluorescent cyanine dyes (Cy 3 or Cy5) using an indirect labeling method. Amino allyl-dUTP is incorporated during reverse transcription and then post-coupled with a NHS-esther mono-reactive cyanine dye. This indirect labeling method allows us to avoid one common form of false positive resulting from the fact that the reverse transcriptase enzyme differentially utilizes the Cy5 and Cy3 dyes. The two labeled cDNA samples are then combined and hybridized on a single microarray slide overnight using a standard formamide-based RNA hybridization protocol (see Protocols toggle on website). The following morning the slide is washed, dried, and scanned using the Axon 4000a scanner. The GenePix® software on the Axon 4000a scanner, after simultaneous laser scanning dual excitation/emission capturing the Cy5 and Cy3 channels, creates a merged 16-bit TIFF image, finds the spots, performs a background subtraction/signal to noise analysis, and then generates a .gpr file containing a variety of parameters concerning each individual spot.
GSM241979 Pituitary 4day(L2);biological replicate Strain: C57BL/6, Gender: male, Tissue: pituitary;Strain: C57BL/6, Gender: male, Tissue: pituitary DNA microarrays were made at the SCCPRR Gene Array Facility (Department of Pharmacology, University of Washington) using the Programs for Genomic Applications (PGA) mouse 70mer oligo library generated at Massachusetts General Hospital (Boston, MA). The mRNAs to be compared are labeled with two different fluorescent cyanine dyes (Cy 3 or Cy5) using an indirect labeling method. Amino allyl-dUTP is incorporated during reverse transcription and then post-coupled with a NHS-esther mono-reactive cyanine dye. This indirect labeling method allows us to avoid one common form of false positive resulting from the fact that the reverse transcriptase enzyme differentially utilizes the Cy5 and Cy3 dyes. The two labeled cDNA samples are then combined and hybridized on a single microarray slide overnight using a standard formamide-based RNA hybridization protocol (see Protocols toggle on website). The following morning the slide is washed, dried, and scanned using the Axon 4000a scanner. The GenePix® software on the Axon 4000a scanner, after simultaneous laser scanning dual excitation/emission capturing the Cy5 and Cy3 channels, creates a merged 16-bit TIFF image, finds the spots, performs a background subtraction/signal to noise analysis, and then generates a .gpr file containing a variety of parameters concerning each individual spot.
GSM241980 Pituitary 4day(L2s);biological replicate_dye swap Strain: C57BL/6, Gender: male, Tissue: pituitary;Strain: C57BL/6, Gender: male, Tissue: pituitary DNA microarrays were made at the SCCPRR Gene Array Facility (Department of Pharmacology, University of Washington) using the Programs for Genomic Applications (PGA) mouse 70mer oligo library generated at Massachusetts General Hospital (Boston, MA). The mRNAs to be compared are labeled with two different fluorescent cyanine dyes (Cy 3 or Cy5) using an indirect labeling method. Amino allyl-dUTP is incorporated during reverse transcription and then post-coupled with a NHS-esther mono-reactive cyanine dye. This indirect labeling method allows us to avoid one common form of false positive resulting from the fact that the reverse transcriptase enzyme differentially utilizes the Cy5 and Cy3 dyes. The two labeled cDNA samples are then combined and hybridized on a single microarray slide overnight using a standard formamide-based RNA hybridization protocol (see Protocols toggle on website). The following morning the slide is washed, dried, and scanned using the Axon 4000a scanner. The GenePix® software on the Axon 4000a scanner, after simultaneous laser scanning dual excitation/emission capturing the Cy5 and Cy3 channels, creates a merged 16-bit TIFF image, finds the spots, performs a background subtraction/signal to noise analysis, and then generates a .gpr file containing a variety of parameters concerning each individual spot.
Platform GPL6050: SCCPRR_Mouse_19549_v1.1
Indicator
Pip5k, Flt1, Clk4, Csnk1, Ptp, Ppp2r2a, Cdcn1a, Rbbp7, Rhoa, Rab4a, Atp5j2, tp5e, Mt1, Lcn2, Hspa8, Hspa5, Scg2, Psma2, Npy, Npyr Y1
AgRP, CPE, Lepr, Peptide YY, Adiponutrin, Pomc1, CART, RIKEN cDNA 5730513H21, Unknown (RIKEN cDNA 3110001K13, AgRP, Npy, proSAAS, ACP homolog, AgRP
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