Northern hybridization

B-raf, a member of the raf gene family of serine/threonine kinases, is expressed as two major transcripts of 4.0 kb and 2.6 kb in the mouse testis (Wadewitz et al., 1993). B-raf expression is limited to the germ cells and is particularly abundant in early spermatids. Northern hybridization analysis revealed that the two B-ra/transcripts are expressed in a stage-specific manner. Low levels of the 4.0-kb transcript are first... 

Adapted cells were harvested at 0 h, 1 h, and 3 h, and total RNA was isolated as described in Materials and Methods. Northern hybridization with the ars specific probe (A) and the hydA specific probe (B) are shown. 

Northern hybridization of the clone to mRNA from human cells revealed the presence of two mRNA species of 0.7 and 0.5 kb. The two mRNAs were found to code for the same polypeptide chain and were transcribed from the same gene. The major 0.7-kb mRNA was much more abundant than the 0.5-kb one. The larger mRNA contained 222 additional nucleotides at the 3 -polyadenylated terminus, and both species had multiple 5 ends. 

The human Y1 mRNA is about 3.5-4 kb and was detected in the neuroblastoma cell line SK-N-MC by Northern hybridization (Larhammar et al., 1992). An in situ hybridization study with a riboprobe reported widespread distribution in human fetal and adult organs, e.g. colon, kidney, heart, placenta and adrenal (Wharton etal., 1993). However, a Northern hybridization in the same study detected a single mRNA of only 2.2 kb suggesting that the specificity of the probe for Y1 mRNA may be questioned. Curiously, in the same study the size for NPY mRNA was reported as 3.3 kb, which disagrees with the size seen in a human pheochromocytoma (0.8 kb) as well as the size of the human cDNA clone which ends with a poly(A) tract (Minth et al., 1984). 

The VDE mRNA transcript level was analyzed in market romaine lettuce by northern hybridization. The younger leaf tissues (yellow leaves and rapidly expanding green leaves) had low levels of transcript... 

Northern hybridization is used to locate the RNA species of interest. The RNA is immobilized and fixed on a membrane. Hybridization with specific probes is then performed under optimized conditions and the obtained image of the distribution of the tightly bound probe on the membrane is read. At the end of the procedure, the probe may be stripped from the membrane and the membrane could be used for another hybridization. 

The function of the nuclear encoded PSI subunits is still unknown. Mutants affected in the synthesis of these subunits could provide valuable insights into this problem. We have therefore screened 58 PSI mutants by Northern hybridization using the cDNAs of P21, P28, P30, P35 and P37 as probes. No significant change in the transcript levels of any of these subunits was found. A similar approach with nuclear PSII mutants has identified mutations affecting the transcription and/or stabilization of the mRNAs of the PSII OEEl and 0EE2 proteins (17,18). One possibility is that the absence of one of these nuclear encoded PSI subunits still allows for partial PSI activity. Mutants of this sort would be leaky in contrast to the mutants screened which were all completely deficient in PSI activity. Another possibility is that some of the mutants are affected at the translational level. Antibodies against the individual polypeptides are required to test this hypothesis. 

Transcripts from this RNA polymerase subunit operon have been characterized by Northern hybridization. rpoB, rpoCl, and rpoC2 gene-specific probes all hybridize to... 

The relative amounts of polypeptides in the cotyledons were determined by a Western blotting analysis. The relative synthesis rates of proteins were determined by in vivo labeling with (135s] Met and the immunoprecipitation method. The relative amounts of hybridizable mRNA were determined by a Northern hybridization method. 

Tomato (Lycopersicon esculenturn cv Platense) chloroplasts (9) and chromoplasts (3) were isolated by previously published procedures. Pt DNA and total RNA were prepared as described in Ref. 3 and 9, respectively. Conditions for agarose electrophoresis of DNA restriction fragments, electrophoresis of RNA in formaldehyde/formamide/agarose gels Southern and Northern hybridizations were esentially those of Ref. 10. 

FIGURE 3. (right) Transcription of the cloned genomic region as analyzed by Northern hybridization. RNA (10 pg) was electrophoresed on a 1% agarose gel containing 0.7 M formaldehyde, tr s-ferred to Zeta-Probe (BioRad) and hybridized with the P-labeled 0.87 kb PstI fragment. The RNA was prepared from cells exposed to I0W-CO2 for 2 hours. 

The cyanobacterial strain Anabaena variabilis PCC 29413 was maintained as described (3) on solid and liquid media supplemented with 10 mM HEPES pH 8.0. High CO2 cultures were stirred and bubbled with 2-5 % CO2 in air while low CO2 cultures were aerated with air alone (0.035 % CO2). The bacterial strains E.. coli HBlOl and JMlOl were maintained as described (4). Cyanobacterial genomic DNA and bacterial plasmid DNA was isolated essentially as described (5) with some modifications. Southern hybridization of restriction endonuclease fragments was performed with nitrocellulose membranes using P-labelled heterologous probes according to the manufacturer s specifications (S S). Total RNA isolation from Anabaena cells was performed as described (6) with some modifications. Northern hybridization was... 

In contrast, there is data supporting that p53 is not a transcriptional activator of p21 in mES cells. Aladjem et al. did not detect p21 expression by immunoblots, immunofluorescence or northern hybridization analysis (Aladjem et al. 1998). This is in agreement with earlier findings of Savatier et al. that did not detect p21 RNA or proteins (Savatier et al. 1996). 

As estimated from the foregoing profile, this gene is transcribed in a mono-cistronic manner without any peripheral genes on the genome, which was confirmed by only one band with a cZpB-targeting probe on Northern hybridization. 

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