Quantitative interference microscopy of polytene chromosomes. I. Cytophysical studies on refractive index and dry mass concentration.

Abstract Quantitative studies on the cytophysical properties of the polytene chromosomes of the salivary gland (first, second, and third instar) and Malpighian tubules (third instar) of Drosophila melanogaster were undertaken by interferometry. Reproducibility of refractive indices (RI) was 0.001 ± 0.0005 units. Isolation of nuclei in aqueous media caused a decrease in dry mass of nuclear components, but isolation in a fluoro-carbon polymer did not reduce the components. During development the dry mass increased in the bands and interbands, but decreased in the nucleoplasm and nucleoli. The apparent nucleolar loss in solids was due to vacuole formation because the vacuole-free zones showed no decrease in RI. For third instar larvae the total percent of dry mass in the nucleoli, bands, and interbands was about 31, 22, and 14, respectively. Band and interband regions were heterogeneous. Proteolytic enzymes (pepsin and trypsin) removed protein from the band and interband regions. Cold shock of larvae decreased dry mass of nucleoli and nucleoplasm, but droplets (possibly protein) appeared near the chromosomes. DNase on fixed cells reduced the material in the bands and interbands. RNase was effective in removing material from bands and nucleoli in living and fixed cells, but no change was detectable in interbands. These enzyme data show that of the band material, about 6 % was DNA and 5% was RNA, while the interbands were only 1 % DNA. The results of chemical extractions (TCA and cold perchloric acid) compare well with these percentages.

[1]  T. Caspersson,et al.  A scanning interference microscope arrangement. , 1954, Experimental cell research.

[2]  K. Ross.  QUANTITATIVE STUDIES ON LIVING CELL INCLUSIONS , 1962 .

[3]  R. W. Rasch.  The productionof specific chromosome alterations with deoxyribonuclease. , 1962, Experimental cell research.

[4]  H. Ragetli,et al.  SOME CONDITIONS AFFECTING THE INTRACELLULAR ARRANGEMENT AND CONCENTRATION OF TOBACCO MOSAIC VIRUS PARTICLES IN LOCAL LESIONS , 1967, The Journal of cell biology.

[5]  R. M. Welch,et al.  APPLICATION OF NAPHTHOL YELLOW S CYTOPHOTOMETRY IN DEOXYRIBONUCLEIC ACID AND PROTEIN DETERMINATIONS ON LARVAL MATERIAL OF D. MELANOGASTER , 1963 .

[6]  H. G. Davies,et al.  Interference Microscopy and Mass Determination , 1952, Nature.

[7]  M. Lezzi.  Die wirkung von DNase auf isolierte Polytän-Chromosomen , 1965 .

[8]  L. VOGT-KOHNE,et al.  [Cytochemical investigations on the Balbiani rings of the 4. salivary gland chromosome of Chironomus tentans]. , 1963, Chromosoma.

[9]  B. Kaufmann.  Cytochemical studies of the action of trypsin: III. The course of deformation of salivary-gland chromosomes , 1953 .

[10]  R. E. Meyer,et al.  Osmotic Stress: Effects of Its Application to a Portion of Wheat Root Systems , 1964, Science.

[11]  M. Lezzi.  [Cytochemical studies of puffs of isolated salivary-gland-chromosomes of Chironomus]. , 1967, Chromosoma.

[12]  Joseph G. Hoffman,et al.  Physical Techniques in Biological Research , 1963 .

[13]  G. Bahr,et al.  The submicroscopic structure of Balbiani-ring. , 1954, Experimental cell research.

[14]  D. Wolstenholme.  Direct evidence for the presence of DNA in interbands of Drosophila salivary gland chromosomes. , 1966, Genetics.

[15]  A. Pollister,et al.  An autoradiographic study of RNA synthesis in isolated salivary glands of Drosophila hydei. II. Interferometric studies , 1965, Journal of morphology.

[16]  B. Ephrussi,et al.  A Technique of Transplantation for Drosophila , 1936, The American Naturalist.

[17]  M. Kopac.  SECTION OF BIOLOGY: RECENT DEVELOPMENTS IN CELLULAR MICRURGY* , 1955 .

[18]  A. Pearse.  Histochemistry: Theoretical and Applied , 1953 .

[19]  M. Chiga,et al.  MICROBODIES IN EXPERIMENTALLY ALTERED CELLS , 1970, The Journal of cell biology.

[20]  D. Mazia.  FIBER PROTEIN STRUCTURE IN CHROMOSOMES AND RELATED INVESTIGATIONS ON PROTEIN FIBERS , 1950 .

[21]  J S Paul,et al.  Quantitative interference microscopy of polytene chromosomes. II. Cytochemical studies via ultramicrospectrophotometry. , 1970, Experimental cell research.

[22]  G. Wied,et al.  Introduction to quantitative cytochemistry , 1967 .