A structure of potentially active and inactive genes of chicken erythrocyte chromatin upon decondensation.

In the presence of 3 mM MgCl2 DNase I cleavage of bulk, globin and ovalbumin gene chromatin in chicken erythrocyte nuclei generates fragments which are multiples of a double-nucleosome repeat. However, in addition to the dinucleosomal periodicity beta-globin gene chromatin was fragmented into multiples of a 100 b.p. interval which is characteristic for partially unfolded chromatin. This distinction correlates with higher sensitivity of beta-globin domain to DNase I and DNase II as compared to the inactive ovalbumin gene. At 0.7 mM MgCl2 where these DNases fragment bulk chromatin into series of fragments with a 100 b.p. interval, the difference in digestibility of the investigated genes is dramatically decreased. When chromatin has been decondensed by incubation of nuclei in 10 mM Tris-buffer, DNase II generates a typical nucleosomal repeat, and the differential nuclease sensitivity of the analyzed genes is not observed. The data suggest that higher nuclease sensitivity of potentially active genes is due to irregularities in higher order chromatin structure

Production of monoclonal antibodies against recombinant human zona pellucida glycoproteins: utility in immunolocalization of respective zona proteins in ovarian follicles

The zona pellucida (ZP) glycoproteins play an important role in oocyte development and gamete biology. To analyze their expression in follicles during various developmental stages, murine monoclonal antibodies (MAbs) were generated against the baculovirus-expressed recombinant human ZP2, ZP3 and ZP4. A panel of MAbs specific for the respective zona protein in ELISA and Western blot, and devoid of cross-reaction with other zona proteins was selected. Immunohistochemistry has shown that ZP2 MAb, MA-1620, did not react with oocytes in resting primordial follicles but showed reactivity with degenerating oocytes in primordial follicles undergoing atresia, and with oocytes in growing and antral follicles. Three MAbs against ZP3 did not react with oocytes in primordial follicles, but reacted only with oocytes in growing and antral follicles. Out of four MAbs against ZP4, three MAbs reacted with oocytes in primordial, growing and antral follicles. No reactivity of these MAbs with other ovarian cell types and other tissues studied (endometrium, uterine cervix, fallopian tubes and kidney) was detected except for a strong reactivity of ZP2 MA-1620 with epithelial cells of the uterine ectocervix or endometrium in some samples investigated. Altogether, these studies document generation of MAbs exhibiting high specificity for human zona proteins, which will be useful reagents to study their immunobiology