H. Aviv

A comparison of globin genes in duck reticulocytes and liver cells

Abstract Synthetic duck globin DNA has been used to compare the number of genes corresponding to globin in duck reticulocytes and liver cells. The results indicate that the number of globin genes does not differ significantly in these organs and suggest, therefore, that neither amplification nor extensive reiteration of these genes occurs as these two cell types become committed to divergent lines of development.

Protein synthesis directed by encephalomyocarditis virus RNA II. The in vitro synthesis of high molecular weight proteins and elements of the viral capsid

Abstract RNA from the encephalomyocarditis virus directs the cell-free synthesis of several discrete, high molecular weight proteins. The largest of these have molecular weights of approximately 110,000, 82,000, 73,000, 61,000 and 44,000 Daltons. In addition, tryptic digestion of the in vitro products gives rise to a number of peptides corresponding to those derived from the viral capsid. The data suggest that approximately one-third of the information encoded by the EMC genome is translated in vitro as a single polypeptide chain, that this translation proceeds in an appropriate phase, and that portions of the genome corresponding to structural proteins of the virus are translated.

Characteristics of rabbit globin mRNA purification by oligo(dT) cellulose chromatography

Abstract We have purified rabbit globin mRNA using oligo(dT)-cellulose and sucrose gradient centrifugation. Both α- and β-globulin mRNA molecules behave heterogeneously with respect to their elution properties during chromatography on oligo(dT)-cellulose. Those fractions eluted at the lowest ionic strength are most active in directing cell-free globin biosynthesis. By making use of hybridization with synthetic [3H]DNA complementary to globin mRNA, we have shown that this technique can be used to quantitate the extent of mRNA purification. Thus, globin mRNA is approximately 90-fold purified from reticulocyte polysomal RNA and originally constituted slightly more than 1% of the polysomal RNA. Since more than 98% of the globin mRNA sequences are bound to oligo(dT)-cellulose, we suggest that most polysomal globin mRNAs contain a poly (A)-rich region and that this region is not of uniform length nor preponderately associated with either the α- or β-globin mRNAs. In addition, we observe that the 9S globin mRNA most resistant to dissociation from oligo (dT)-cellulose is most active in directing globin biosynthesis.

Characterization of synthetic rabbit globin DNA

Abstract Synthetic DNA (cDNA) complementary to purified mRNA has already proven to be a useful hybridization probe for the quantitation of specific genes and for the detection of specific mRNAs in eukaryotic organisms. In view of its central role in such studies, it seemed necessary to characterize further one such cDNA and to determine the value of established techniques for detecting DNA-RNA hybrids when cDNA is used as a probe. We, therefore, synthesized cDNA complementary to purified rabbit globin mRNA using avian myeloblastosis virus reverse transcriptase. Equilibrium centrifugation in cesium chloride and nearest-neighbor analysis show this cDNA to be heteropolymeric, with a G,C content of 46–48%. Its base composition and nearest-neighbor analysis reflect the expected high frequency of TpT doublets, but also a relatively high proportion of GpG doublets. The cDNA contains at least one T-rich region of sufficient length to form a hybrid with poly (A). Hybridization involving this region is not detected when S i nuclease is used to detect DNA-RNA hybrids. However, this interaction does affect results obtained using hydroxyapatite chromatography. Thus, rabbit globin cDNA exhibits stringent hybridization specificity when assayed using S 1 nuclease, annealing efficiently to rabbit globin mRNA, and not to ribosomal, transfer, or nonglobin mRNAs.

DNA Complementary to Globin and Immunoglobulin mRNA: A Probe to Study Gene Expression

During the development of an embryo, various processes occur which induce the formation of differentiated tissues. Some of these processes can be observed at a molecular level, for example, myosin is synthesized in muscle cells, crystaline in lens tissue, keratin in skin, immunoglobulin in lymphocytes and hemoglobin in reticulocytes (1,2).