The sequence of Japanese quail ovalbumin cDNA (original) (raw)
Related papers
Biochemistry, 1975
Preparation of milligram amounts of purified ovalbumin mRNA was accomplished by a sequential combination of precise sizing techniques with the selective purification of the poly(A) containing RNA by either affinity chromatography or adsorption to nitrocellulose filters. Several new techniques were applied to the purification of ovalbumin mRNA including Sepharose 4B chromatography and agarose gel electrophoresis in the presence of 6 M urea at pH 3.5. All the procedures used were adapted on a preparative sacle to the fractionation of large quantities of RNA. The purity of the ovalbumin mRNA was assessed by several independent criteria. (1) Purified ovalbumin mRNA migrated as a single band during both agarose-urea and formamide-polyacrylamide gel electrophoresis at pH 3.5 and 7.4, respectively. A single absorbance peak containing all of the ovalbumin mRNA activity was also found using linear formamide-sucrose gradients. (2) Determination of both total mRNA activity and ovalbumin mRNA activity in the wheat germ cell-free translation assay revealed that 92% of the total peptides synthesized were specifically immunoprecipitable with an ovalbumin antiserum. (3) Analysis of the total peptides synthesizied in the wheat germ assay by sodium dodecyl sulfate polyacrylamide gel electrophoresis demonstrated the presence of a single radioactive peak that corresponded exactly to a specifically immunoprecipitable ovalbumin standard. Thus, based on these observations ovalbumin mRNA appears to be greater than 95% pure. A preliminary estimation of the molecular weight of purified ovalbumin mRNA by formamide-containing sucrose gradients yielded a value of 520,000 or approximately 1600 nucleotides. This value was considerably less than the value of 900,000 obtained by gel electrophoresis under denaturing conditions. Analysis of the poly(A) content by a hybridization assay with (3H)poly(U) revealed the presence of a poly(A) region containing approximately 70 adenosine residues. Thus, the size of the ovalbumin mRNA is considerably greater than that required to code for a protein of 387 amino acids. The availability of large quantities of purified ovalbumin mRNA should now permit a more thorough analysis of its physical and chemical properties.
The Complete Amino-Acid Sequence of Hen Ovalbumin
European Journal of Biochemistry, 1981
The complete amino acid sequence of hen ovalbumin, comprising 385 residues, has been determined. The sequence was deduced from the 17 cyanogen bromide fragments and from peptides derived by digestion with a number of proteolytic enzymes. The molecular weight of the polypeptide chain of ovalbumin is 42699.
Nucleic Acids Research, 1980
We have examined homologous fragments of DNA cloned from two different tissues for changes in the DNA sequence which might be related to tissue specific gene expression. The 5' end of the chicken ovalbumin gene was cloned from oviduct or erythrocyte DNA using cosmids as vectors. We have compared the two clones obtained by restriction enzyme digestions, analysis of heteroduplexes by electron microscopy or S1 nuclease digestion and by DNA sequencing. Our results show that whereas no alteration occured in the region of the gene assumed to be of importance for the control of transcription, a 4 nucleotide deletion/insertion was detected in the first intron of the ovalbumin gene.
Age-Related Changes in the Ovalbumin Gene of the Japanese Quail
Biochemical and Biophysical Research Communications, 1996
Northern hybridization studies showed that the level of ovalbumin mRNA decreases in the oviduct of the Japanese quail after adulthood. In order to find out if this is due to changes in the conformation of the chromatin containing the promoter region of the gene, nuclei of the oviduct of young, adult and old birds were digested by DNaseI and micrococcal nuclease (MNase). Southern hybridization with the labelled promoter showed that this region is less sensitive to the two enzymes in the old birds. Both the endonucleases recognized the same hypersensitive sites. The results indicate that the chromatin containing the promoter is present in an open conformation in adult birds. Gel mobility shift assay using a 20-mer dsDNA containing the CAAT-box and nuclear extract of oviduct shows the presence of trans-acting factors that bind to this region. The levels of these factors are lower in old birds. This may be the reason for the lower expression of the gene in old birds. ᭧ 1996 Academic Press, Inc.
Preferential transcription of the ovalbumin gene in isolated hen oviduct nuclei by RNA polymerase B
Proceedings of the National Academy of Sciences, 1978
The synthesis of ovalbumin mRNA sequences was studied in isolated nuclei from hen oviduct. Two different methods of analysis were used to distinguish in vitro synthesized from preexisting mRNA sequences: (i) Mercurated ribonucleotides were used for in vitro RNA synthesis, and the newly synthesized RNA was purified by chromatography on sulfhydryl-agarose and hybridized to radioactive ovalbumin cDNA. (ii) [3H]UTP was used to label the in vitro synthesized RNA. Hybridization to unlabeled mercurated cDNA, RNase A digestion, and subsequent purification of the hybrids on SH-agarose allowed the quantitation of newly synthesized ovalbumin mRNA sequences. Approximately 0.1% of the newly synthesized RNA was identified as ovalbumin RNA by both methods. The synthesis of ovalbumin RNA progressed during the incubation of nuclei and was sensitive to actinomycin D and low concentrations of alpha-amanitin. The preferential in vitro transcription of the ovalbumin gene (1000-fold over random transcrip...
The Journal of biological chemistry, 1978
A scheme is presented for cloning a double-stranded cDNA molecule that codes for a portion of chicken preproalbumin. This method, which does not require pure mRNA or cDNA, has widespread applicability. Chicken preproalbumin was identified as a Mr = 72,000 polypeptide by immunoprecipitation of proteins synehesized in a wheat germ cell-free translation system from total, guanidine.HCl-extracted, rooster liver RNA. After removal of the bulk of the ribosomal RNA by poly(U)-Sephadex G-10 chromatography, albumin mRNA was enriched approximately 2-fold by centrifugation through low salt, isokinetic sucrose gradients, until it represented about 30% of the mRNA sequences present. Double-stranded cDNA prepared from this mRNA was then inserted into the Pst 1 site of the plasmid PBR322 by the "G-C tailing" technique and the recombinant DNA was used to transform Echerichia coli stran X1776. Transformants containing putative albumin DNA sequences were identified by colony hybridization w...
Comparative Biochemistry and Physiology Part C: Pharmacology, Toxicology and Endocrinology - COMP BIOCHEM PHYSIOL C PHARMACOL TOXICOL ENDOCRINOL, 1995
The effects of estrogen, dexamethasone, insulin-like growth factor-I (IGF-I), and transferrin on the messenger RNA (mRNA) contents of ovalbumin and conalbumin in primary cultures of quail oviduct cells were investigated. In the absence of one of the above hormones or factors, a decrease in ovalbumin mRNA was prominent. In particular, removal of IGF-I and transferrin caused a significant effect. Studies using a combination of estrogen, dexamethasone, IGF-I and transferrin indicated that IGF-I cooperates with estrogen or dexamethasone and transferrin works with dexamethasone. Specifically, IGF-I enhanced ovalbumin synthesis or increased cellular ovalbumin mRNA content depending on its concentration in the medium in the presence of estrogen. However, the effects of estrogen, dexamethasone, IGF-I, and transferrin were not similarly observed with conalbumin mRNA. These results show that ovalbumin synthesis is controlled by estrogen or glucocorticoid with IGF-I or transferrin and that cel...