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Biomedical subjects

C L Hew

Publications and source records attributed to C L Hew.

106 records · Page 6Linked to original sources
PubMed1978

Cell-free synthesis of cod preproinsulin.

Poly(A)-containing ribonucleic acid from cod islet greatly stimulated the incorporation of radioactive amino acids into proteins when assayed in a wheat germ translation system. The translation products were examined by specific immunoprecipitation with guinea pig anti cod insulin antibodies and by extraction with acid--ethanol. These measurements revealed at least a fivefold increase in incorporation of labelled amino acid over the nonprogrammed system. Sodium dodecyl sulfate disc gel electrophoresis and gel filtration chromatography showed a product of molecular weight 12 500, a size considerably larger than cod proinsulin (9000). It is concluded that cod proinsulin is synthesized via a larger precursor, preproinsulin.

Animals↗
PubMed1975

Translation of messenger ribonucleic acid from isolated pancreatic islets and human insulinomas.

RNA preparations from isolated rat pancreatic islets and from human insulinomas were injected into oocytes of Xenopus laevis which were then incubated with [3H]leucine. Acid-ethanol extracts of the oocytes were immunoprecipitated with anti-insulin serum using the double antibody technique. Sodium dodecyl sulfate disc gel electrophoresis of the immunoprecipitates showed the presence of an insulin-displaceable immunoreactive material with a molecular weight of about 18,000 in extracts from oocytes injected with RNA of 9-11 S. This immunoreactive product was not detected in extracts from oocytes injected with buffer or 4-8S RNA or RNA heavier than 11 S. These observations suggest the involvement of a precursor larger than proinsulin in the biosynthesis of insulin.

Adenoma, Islet Cell↗
PubMedDate not supplied

Development of an all-fish gene cassette for gene transfer in aquaculture.

To develop an all-fish gene cassette suitable for gene transfer in aquaculture, the antifreeze protein (AFP) gene promoter from the ocean pout (Macrozoarces americanus) was analyzed for its ability to direct exogenous gene expression both in vitro and in vivo. The ocean pout AFP (opAFP) gene promoter fused to the bacterial chloramphenicol acetyltransferase (CAT) was functionally analyzed in two fish cell lines and in Japanese medaka embryos. The opAFP gene promoter was active in these systems, as demonstrated by the transient expression of CAT activity. These results suggest that the opAFP gene promoter is useful for many other gene transfer experiments. To facilitate use of the opAFP gene promoter as a common and versatile vehicle for fish gene transfers, an expression vector, opAFP-V, was constructed by linking the 2.1-kb opAFP gene promoter, the 63-bp opAFP gene 5' untranslated sequence, and the 1.2-kb opAFP gene 3' sequence by two unique restriction sites, Bg/II and HpaI, respectively. Thus, genes of interest can be inserted into either the Bg/II site or the HpaI site depending on the length of their 5' untranslated sequence. The complete DNA sequence of opAFP-V was determined to facilitate future detailed analysis of integration and expression of the transgene.

Animals↗
PubMedDate not supplied

Antifreeze protein gene transfer in Atlantic salmon.

Salmonids freeze to death if they come into contact with ice. Many marine fish species that inhabit icy sea waters synthesize antifreeze proteins (AFP) to protect them from freezing. Production of stable lines of freeze-resistant salmon and other species would greatly facilitate development of sea-pen aquaculture in many regions. We successfully introduced winter flounder AFP genes into Atlantic salmon. Research to date indicates stable genomic integration and low levels of expression of winter flounder AFP genes in a small number (approximately 3%) of salmon developed from microinjected eggs. Inheritance of the AFP gene by offspring (F1) from crosses between transgenic and wild-type salmon revealed that the transgenic flounders (F0) were germ-line mosaics. Low levels of AFP precursors could be detected in the blood of all these transgenic offspring (F1). Approximately 50% of the progeny produced by crosses between transgenic F1 and wild-types contained the AFP genes. These results demonstrate that stable germ-line transformed Atlantic salmon can be produced.

Animals↗
PubMedDate not supplied

Isolation and characterization of somatolactin genes from two cold water marine teleosts, lumpfish (Cyclopterus lumpus) and halibut (Hippoglossus hippoglossus).

Somatolactin (SL) is a newly discovered pituitary hormone structurally related to both growth hormone and prolactin. We isolated the SL gene from two cold water marine teleosts, lumpfish (Cyclopterus lumpus) and halibut (Hippoglossus hippoglossus). The SL genes for lumpfish were isolated from a complementary DNA (cDNA) library by random gene tagging and sequencing. Two full-size clones of 1.2 (lfSL-6) and 1.6 (lfSL-9) kilobases (kb) were characterized. The SL genes for halibut were identified from its cDNA library using lumpfish SL gene as a probe. Two full-size SL clones of 1.1 and 1.6 kb were identified in halibut. All these clones encoded full-size SL (229 amino acids for lumpfish SL, 230 amino acids for halibut SL), including 7 cysteines and two and three potential glycosylation sites in halibut and lumpfish, respectively. The differences in length of these clones are attributed to their differences in 3' untranslated sequences. Although the halibut SL genes contain the consesus polyadenylation signal AATAAA, the lumpfish SL gene has the ATTAAA sequence in the larger clone (lfSL-9). None of the consensus or the alternative polyadenylation signals were found in the shorter clone (lfSL-6). Protein homology and DNA sequence alignments of SL genes from all five known marine fishes suggest that the SL gene sequence is highly conserved.

Amino Acid Sequence↗