A digoxigenin-labeled nonradioactive detection system was used
to screen a zebrafish cDNA library for p53-like and ras-like genes.
One clone was isolated and identified as an incomplete p53-like
gene. The insert size of this clone is 1777 bp, which encodes part of
evolutionarily conserved region II and all of regions III, IV, and V. A
magnetically enriched whole zebrafish cDNA library was constructed
to enhance possible recovery of ras-like genes in zebrafish. One
clone, termed Zras-Bl, carried an insert of 2592 bp with an open
reading frame encoding a 188 amino acid residue ras p21 protein.
Based on total protein sequence, this expressed zebrafish ras p21 is
most closely related to human N-ras (91% homology), with lesser
homology to Ha-ras (84%) and Ki-ras (85%). Preliminary partial
sequence data obtained by genomic and reverase transcriptasepolymerase
chain reaction (RT-PCR) screening indicate the presence
of at least one additional expressed ras gene in zebrafish.
The tumorigenicity and Ki-ras mutational properties of dietary
7,12-dimethylbenz[a]anthracene (DMBA) and dibenzo[a,l]pyrene (DBP) were compared in rainbow trout. Both chemicals elicited
predominantly 12(1)G->A and 12(2)G->T mutations in trout liver
tumors. Two {12(1)G->T and 12(2)G->T} and one {12(1)G->A and
12(2)G->T} double mutation were also observed in DBP livers
tumors, but not in DMBA liver tumors. Some stomach tumors from
both chemicals exhibited so much DNA degradation that routine PCR
amplification was not possible. Among sixteen DMBA stomach
tumors with intact DNA, no Ki-ras mutations were found. Of sixteen
DBP stomach tumors examined, one had 12(1)G->A and two had
13(1)G->C mutations. The observed G->T transversions are
compatible with apurinic mutagenesis driven by unstable DNA
adducts arising from one-electron oxidation, but this is not true for
the major G->A transitions or G->C transversions and rare double
mutations found in this study. The low sensitivity of direct
sequencing may limit the frequency of ras mutant detection in this
study. / Graduation date: 1996
| Identifer | oai:union.ndltd.org:ORGSU/oai:ir.library.oregonstate.edu:1957/27187 |
| Date | 08 August 1995 |
| Creators | Cheng, Ronshan |
| Contributors | Bailey, George S. |
| Source Sets | Oregon State University |
| Language | en_US |
| Detected Language | English |
| Type | Thesis/Dissertation |
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