Radomir Crkvenjakov

Variant chromosomal arrangement of adult β-globin genes in rat

Abstract The genomic organization of three haplotypes of β-globin genes was determined to resolve the question of the number of those genes in rat. Haplotype a, found in inbred strain DA, has three genes or pseudogenes, while haplotypes b, found in AO, Y5 and Wistar strains, and c, found in Wistar strain, have five genes or pseudogenes each. In haplotypes b and c, the first gene is of βmajor type and the remaining four are of βminir type. Partial sequencing of six out of 13 genes shows that duplications of βminor genes are causing polymorphism in a number of genes. Also, in haplotype b two βminor genes have a 6.5-kb intron 2, while in haplotype c only one βminor gene contains such a large intron 2. The three structurally different haplotypes described are not interconvertible by single recombination events. The results indicate that the rat has the highest number of adult β-globin genes found in mammals so far.

Agrobacterium-mediated transformation and plant regeneration of buckwheat (Fagopyrum esculentum Moench.)

Genetic transformation of buckwheat (Fagopyrum esculentum Moench.) and regeneration of transgenic plants were obtained by using Agrobacterium tumefaciens strains as vectors. Buckwheat cotyledons were excised from imbibed seeds, co-cultivated with A. tumefaciens and subjected to previously reported protocols for callus and shoot regeneration. The transformation with oncogenic strains was confirmed by opine and DNA analyses of tumour tissue extracts. Plants were regenerated on cotyledon fragments incubated with strain A281, harboring pGA472, which carries the neomycin phosphotransferase II gene for kanamycin resistance. The transformation of resistant shoot clones was confirmed by NPTII enzyme assay and DNA hybridization. A large number of transformed shoots were rooted and fertile plantlets were raised in the greenhouse. Transgenic plants comprised pin and thrum clones, which were allowed to cross-pollinate. In about 180 R2 seeds tested for kanamycin resistance, the ratio of resistant to sensitive seedlings was roughly 3:1.

A pool of nonpolysomal globin mRNAs in globin deficient reticulocytes of the anemic Belgrade rat

Abstract In anemia of the Belgrade rat ( b b ) reticulocytes contain less than half of the normal amount of mRNA for seven adult rat globin chains. cDNA hybridization measurements of the relative sizes of polysomal and nonpolysomal pools of globin mRNAs in these cells show that 45% of all globin mRNA molecules are not used at any given time in protein synthesis. This implies a translational control which ensures a production of globin chains in a correct ratio despite a severe mRNA unbalance.

Molecular Biology in Embryology

Modern work in embryology puts the old controversy between preformation and epigenesis in a new light. The rapidly dividing cells of embryos contain many maternally preformed, high-molecular-weight components, including both enzymes and molecular building blocks to be used long after fertilization. All presently investigated cellular macromolecules, with the exception of DNA, are found in excess in fertilized egg cells, compared to levels in mature somatic cells. Many of these macromolecules, including an interesting population of stable RNA’s, remain constant in total amount through many successive cell divisions. Identical sets of genes in the cells of the early embryo later give rise to different cell types and lineages. Although some intermediate steps in this differentiation process have been identified, the ultimate cause of differential gene expression remains obscure. It is clear, however, that most differentiation is notthe result of loss of nuclear genetic information. Artificially induced changes in the pattern of cell division in some cases create a disorganized embryo, but in others development proceeds normally. The polarity of molecular organization in an egg’s cytoplasm appears to be critical to normal development. Cytoplasmic polarity may in turn result from the orientation of material in the cell nucleus. There seems to be an intrinsic geometry of chromosomes that permits a spatial display of information there to create significant, stable, and essential inhomogeneities and regionalizations of the cytoplasm. These, in turn, determine subsequent cleavage planes as cells divide, and the partitioning of information between daughter cells. The glossary in Chapter 5 will be helpful here also. In addition, for the nonbiological specialist, we offer the following, brief account of the main processes that relate DNA to proteins through RNA and that permit copying of genes during cell division.