, Volume 6, Issue 1, pp 489–521 | Cite as

Autoradiographic and microphotometric studies of desoxyribose nucleic acid during microgametogenesis in lilium longiflorum

  • J. Herbert Taylor
  • Rachel D. McMaster


  1. 1.

    The technique for making autoradiographs with stripping film is described in detail. Methods for administering P32 to plants and for preparation of tissues for autoradiographs are given.

  2. 2.

    Two different methods, enzyme digestion and acid hydrolysis, which were used for separating RNA, DNA, and phosphoprotein in tissue sections, were compared and evaluated. Although either method is valid for quantitative work, precautions are necessary and certain limitations to the quantitative separation of RNA and DNA should be taken into account. More work is needed to properly characterize the products extracted during enzyme digestion and acid hydrolysis.

  3. 3.

    Incorporation of P32 into nuclei of lily anthers was studied by the autoradiographic method at all stages from premeiosis to mature pollen. A quantitative comparison of the amounts of P32 extracted by RNase, DNase, Feulgen hydrolysis and hot TCA showed that incorporation into DNA occurred at relatively short periods during interphase, namely: (1) at preleptotene before meiotic prophase in microsporocytes, (2) at late interphase preceding mitosis in the microspore and (3) during early to mid-interphase in the generative nucleus of the pollen grain (the tube nucleus incorporates P32 slowly during the same period but this P32 was not definitely identified as DNA phosphorus). Two successive mitoses occur in tapetal cells while the microsporocytes are in meiotic prophase. In the interphase preceding each division incorporation of P32 into DNA occurs, but following the second mitosis no further divisions or incorporation was detected. However, during this latter period incorporation into RNA and phosphoprotein reached the highest level detected at any stage.

    During the period of DNA synthesis, incorporation of P32 into RNA was low or absent, but in the interval between the cessation of DNA synthesis and prophase the incorporation of P32 into RNA of both nucleus and cytoplasm reached its peak.

  4. 4.

    Microphotometric determination of amounts of DNA (Feulgen stain) showed that the amount per nucleus remains constant in all stages, except for doubling during the periods when P32 is incorporated, and the reduction by half at each division of a nucleus. The 2C amount is present in sporogenous nuclei until preleptotene when values intermediate between 2C and 4C are found. By the beginning of leptotene 4C is present in each microsporocyte nucleus. The amount does not change through meiotic prophase. Each nucleus after the first meiotic division contains 2C and each nucleus of the quartet of spores has 1C. The amount per nucleus in microspores doubles toward the end of the long interphase prior to prophase. After the mitosis in the microspore, the generative nucleus attains the 2C level relatively early in interphase. The vegetative nucleus has 1C in the interphase immediately after mitosis, but could not be accurately measured at later stages.

    Tapetal nuclei have the 2C amount of DNA when the microsporocytes are in leptotene. The amount doubles while the microsporocytes are in zygotene and the first mitosis usually occurs while the microsporocytes are in early pachytene. Another doubling occurs during the relatively short interphase and a second mitosis occurs when the microsporocytes are in late pachytene or early diplotene. The 4C class was typical for tapetal nuclei at later stages, but this may, in some cases at least, result from fusion of nuclei in the multinucleate cells during or following the second mitosis.

  5. 5.

    The periods of DNA increase (measured microphotometrically) coincide very closely with the periods of incorporation of P32 into DNA. This finding supports the concept of the stability of the DNA molecule, and indicates that incorporation may be used as a criterion not only of synthesis of DNA, but also of increase in the amount of DNA per nucleus.



Meiotic Prophase Generative Nucleus Early Pachytene Late Pachytene Tube Nucleus 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Alfert, M.: A cytochemical study of oögenesis and cleavage in the mouse. J. Cellul. a. Comp. Physiol.36, 381–406 (1950).Google Scholar
  2. Alfert, M., andH. Swift: Nuclear DNA constancy: a critical evaluation of some exceptions reported byLison andPasteels. Exper. Cell Res.5, 455–460 (1953).Google Scholar
  3. Barnum, C. P., R. A. Huseby andH. Vermund: A time study of the incorporation of radiophosphorus into the nucleic acids and other compounds of a transplanted mouse mammary carcinoma. Cancer Res.13, 880–889 (1953).PubMedGoogle Scholar
  4. Brues, A. M., M. M. Tracy andW. E. Cohn: Nucleic acids of rat liver and hepatoma: their metabolic turnover in relation to growth. J. of Biol. Chem.155, 619–633 (1944).Google Scholar
  5. Bryan, J. H. D.: DNA-protein relations during microsporogenesis ofTradescantia. Chromosoma4, 369–392 (1951).PubMedGoogle Scholar
  6. Cooper, D. C.: The transfer of desoxyribose nucleic acid from the tapetum to the microsporocytes at the onset of meiosis. Amer. Naturalist86, 219–229 (1952).Google Scholar
  7. Erickson, R. O.: Cytological and growth correlations in the flower bud and anther development ofLilium longiflorum. Amer. J. Bot.35, 729–739 (1948).Google Scholar
  8. Hevesy, G., andJ. Ottesen: Rate of formation of nucleic acid in the organs of the rat. Acta physiol. scand. (Stockh.)5, 237–247 (1943).Google Scholar
  9. Howard, A., andS. R. Pelc: Nuclear incorporation of P32 as demonstrated by autoradiographs. Exper. Cell Res.2, 178–187 (1951).Google Scholar
  10. Kaufmann, B. P., H. Gay andM. R. McDonald: Enzymatic degradation of ribonucleoprpteins. Amer. J. Bot.38, 268–275 (1951).Google Scholar
  11. Leblond, C. P., C. E. Stevens andR. Bagorach: Histological localization of newly-formed desoxyribonucleic acid. Science (Lancaster, Pa).108, 531–533 (1948).Google Scholar
  12. Leuchtenberger, C., G. Klein andE. Klein: The estimation of nucleic acids in individual isolated nuclei of Ascites tumors by ultraviolet microspectrophotometry and its comparison with chemical analysis. Cancer Res.12. 277 (1952).Google Scholar
  13. McMaster, Rachel D.: The desoxyribonucleic acid content of nuclei in the sea urchin,Lytechinus variegatus, during development. Anat. Rec.113, 26 (1952). Abstract.Google Scholar
  14. Moses, M. J., andJ. H. Taylor: Desoxypentose nucleic acid synthesis during microsporogenesis inTradescantia. Rec. Genetics Soc. Amer.22, 88–89 (1953). Abstract.Google Scholar
  15. Nelson, P. K.: Ph. D. thesis, Univ. of Wisconsin 1951. Quoted byC. L. Huskins, Nuclear reproduction. Internat. Rev. Cytol.1, 9–26 (1952).Google Scholar
  16. Ogur, M., R. O. Erickson, G. U. Rosen, K. B. Sax andC. Holden: Nucleic acids in relation to cell division inLilium langifloum. Exper. Cell Res.2, 73–89 (1951).Google Scholar
  17. Ogur, M., andG. U. Rosen: The nucleic acids of plant tissues. I. The extraction and estimation of desoxypentose nucleic acid and pentose nucleic acid. Arch. of Biochem.25, 262 to 276 (1950).Google Scholar
  18. Ornstein, L.: The distributional error in microspectrophotometry. Labor. Invest.1, 250–262 (1952).Google Scholar
  19. Pasteels, J., etW. S. Bullough: Augmentation expérimentale de la teneur en DNA des noyaux. (Action de l'huile de crotonsur l'epiderme de l'oreille de souris). Archives de Biol.54, 271–294 (1953).Google Scholar
  20. Pasteels, J., etL. Lison: Etude quantitative de l'acide desoxyribonucléique au cours de la formation du pollen chezTradescantia virginiana. C. r. Acad. Sci. Paris233, 196–199 (1951).PubMedGoogle Scholar
  21. Patau, K.: Absorption microphotometry of irregular shaped objects. Chromosoma5, 341–362 (1952).PubMedGoogle Scholar
  22. Patau, K., u.H. Swift: The DNA-content (Feulgen) of nuclei during mitosis in a root tip of onion. Chromosoma6, 149–169 (1953).PubMedGoogle Scholar
  23. Plaut, W. S.: DNA synthesis in the microsporocytes ofLilium henryi. Hereditas39, 439–444 (1953).Google Scholar
  24. Pollister, A. W.: Photomultiplier apparatus for microspectrophotometry of cells. Labor. Invest.1, 106–114 (1952).Google Scholar
  25. Pollister, A. W., andH. Ris: Cytological nucleoprotein determination in preparations. Cold Spring Harbor Symp. Quant. Biol.12, 157 (1947).Google Scholar
  26. Schrader, F., andC. Leuchtenberger: Variations in the amount of desoxyribose nucleic acid in different tissues ofTradescantia. Proc. Nat. Acad. Sci. U.S.A.35, 464–468 (1949).Google Scholar
  27. Stevens, C. E., R. Daoust andC. P. Leblond: Rate of synthesis of desoxyribonucleic acid and mitotic rate in liver and intestine. J. of Biol. Chem.202, 177–186 (1953).Google Scholar
  28. Stowell, R. E.: Feulgen reaction for thymonucleic acid. Stain Technol.20, 45–46 (1945).Google Scholar
  29. Swift, H.: The constancy of DNA in plant nuclei. Proc. Nat. Acad. Sci. U.S.A.36, 643–654 (1950a).Google Scholar
  30. —: The desoxyribose nucleic acid content of animal nuclei. Physiologic. Zool.23, 169–198 (1950b).Google Scholar
  31. —: Quantitative aspects of nuclear nucleo proteins. Internat. Rev. Cytol.2, 1–76 (1953).Google Scholar
  32. Taylor, J. H.: The duration of differentiation in excised anthers. Amer. J. Bot.37, 137–143 (1950).Google Scholar
  33. —: Autoradiographie detection of incorporation of P32 into chromosomes during meiosis and mitosis. Exper. Cell Res.4, 169–179 (1953).Google Scholar
  34. Walker, P. M. B., andHelen B. Yates: Nuclear components of dividing cells. Proc. Roy. Soc. Lond., Ser. B140, 274–300 (1952).Google Scholar

Copyright information

© Springer-Verlag 1953

Authors and Affiliations

  • J. Herbert Taylor
    • 1
  • Rachel D. McMaster
    • 1
  1. 1.Department of BotanyColumbia UniversityNew York

Personalised recommendations