Northern blot analysis of gene expression in neutrophils: quantitative changes in mRNA levels induced by incubation with opsonized E coli
| Gene symbol . | Transcript encoding . | Relative Northern blot expression . | Differential display expression . | |
|---|---|---|---|---|
| Gene name . | Control . | + E coli . | ||
| IL1B | IL-1, beta | 71 | 2 | 8 |
| IL8 | IL-8 | 16 | 1 | 6 |
| SCYA3 | Small inducible cytokine A3; MIP1A | 41 | 1 | 6 |
| ATP6L | ATPase, H+transporting, lysosomal (vacuolar proton pump) 16 kd | 2.9 | 2 | 3 |
| ITGB2 | Integrin, beta 2 (CD18, LFA-1) | 0.60 | 2 | 1 |
| IL8RB | IL-8 receptor, beta (CXCR2) | 0.21 | 6 | 2 |
| IL8RA | IL-8 receptor, alpha (CDw128a CXCR1) | 0.16 | 4 | 1 |
| NCF1 | Neutrophil cytosolic factor 1 (p47-PHOX) | 0.10 | 6 | 2 |
| Gene symbol . | Transcript encoding . | Relative Northern blot expression . | Differential display expression . | |
|---|---|---|---|---|
| Gene name . | Control . | + E coli . | ||
| IL1B | IL-1, beta | 71 | 2 | 8 |
| IL8 | IL-8 | 16 | 1 | 6 |
| SCYA3 | Small inducible cytokine A3; MIP1A | 41 | 1 | 6 |
| ATP6L | ATPase, H+transporting, lysosomal (vacuolar proton pump) 16 kd | 2.9 | 2 | 3 |
| ITGB2 | Integrin, beta 2 (CD18, LFA-1) | 0.60 | 2 | 1 |
| IL8RB | IL-8 receptor, beta (CXCR2) | 0.21 | 6 | 2 |
| IL8RA | IL-8 receptor, alpha (CDw128a CXCR1) | 0.16 | 4 | 1 |
| NCF1 | Neutrophil cytosolic factor 1 (p47-PHOX) | 0.10 | 6 | 2 |
The relative Northern blot expression is expressed as neutrophils + E coli/control neutrophils. The total RNA of identically treated neutrophils used for either Northern blots or differential display.
RNA from HeLa cells was used as negative control for each measurement. Hybridization probes were cDNA-labeled by random priming of PCR products amplified from neutrophil cDNA. Primer sequences are available on request. Procedures for filter hybridization were previously described.25 Equal loading of lanes was demonstrated by ethidium bromide staining and by rehybridization with a 5.8-kbHindIII restriction fragment of rat 18S ribosomal cDNA.26 Transcript levels in total RNA were quantified by the Phosphorimager System. Data are presented as the means of 2-4 determinations, within 20% of each other. The ratio of expression level of neutrophils incubated with E coli K12 for 2 hours to neutrophils incubated without bacteria for 2 hours is shown. See Figure 1 for the relation between Phosphorimager values and differential display estimates.