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1.
Using β-glucuronidsase the cell walls of tomato and potato primary microsporocytes can be digested. When the resulting protoplasts are exposed to distilled water, they burst, and complete sets of synaptonemal complexes are released to settle on plastic coated slides. After drying and formalin fixation, the synaptonemal complexes can be stained with silver or phosphotungstic acid and observed in the light and/or electron microscope. Silver staining gives better contrast for both light and electron microscopy but stains only lateral elements and kinetochores. Phosphotungstic acid staining gives little or no contrast for light microscopy, but stains both the lateral and central elements of the synaptonemal complex, kinetochores, and structures that are probably recombination nodules for electron microscopy. This technique offers a powerful tool for genome analysis by allowing (1) the determination of relative and absolute lengths of synaptonemal complexes and chromosome arm ratios at pachytene, (2) the analysis of complex patterns of synapsis, and (3) the location of what are probably recombination nodules along the length of synaptonemal complexes.  相似文献   

2.
The distribution of DNA in synaptonemal complexes of rat and mouse spermatocytes, and of chick oocytes was investigated by immunogold electron microscopy. Except for a few specific sites, DNA was not immunolocalized in the space between lateral elements of the complex. Some labeled fibrils connecting the lateral elements with the central element were observed associated with recombination nodules or near them. However, other labeled fibrils in the space between lateral elements did not appear to present any relationship to recombination nodules. The immunocytochemical approaches used here confirmed the presence of significant amounts of DNA in the lateral elements as previously indicated by preferential DNA staining methods. Furthermore, our findings support the view that recombination nodules are the site of chiasma formation.  相似文献   

3.
M I Pigozzi  A J Solari 《Génome》1999,42(2):308-314
Pigeon spermatocytes were processed with a drying-down technique and their synaptonemal complex (SC) complements were analyzed by electron microscopy. The synaptonemal complex karyotype of the macrobivalents shows an excellent correspondence with the mitotic karyotype. The number and distribution of recombination nodules (RNs) were scored in complete nuclei stained with phosphotungstic acid. The average number of RNs per nucleus is 64.7. The number of nodules per bivalent shows a clear linear relationship with SC length in the 10 longest synaptonemal complexes, while the microbivalents usually bear a single RN. The location of RNs has a non-random distribution along the largest synaptonemal complexes, with lower frequencies near kinetochores and higher frequencies toward the telomeres. The ZZ bivalent is the fourth in size and shows free recombination, having on average 3.8 RNs. The mean number of nodules per cell and the mean number of nodules in the largest bivalents show very good agreement with the corresponding number of chiasmata scored in metaphase-I spermatocytes. It is concluded that the recombination nodules provide a good check for reciprocal exchanges in this and other species of birds. Additionally, a new morphology for the recombination nodules is presented, consisting of groups of electron-dense particles measuring 43 nm in diameter.  相似文献   

4.
In most organisms, biological variability rests on the behaviour of the chromosomes in the meiotic context. Despite the importance of meiosis, very little is known about the meiotic behaviour of the Eimeria chromosomes. The aim of the present study is to describe the standard synaptonemal complex karyotype from Eimeria tenella oocyst spreads by electron microscopy. For that purpose, complete sets of pachytene synaptonemal complexes were obtained and the morphological pachytene karyotype was determined. The authors used a previously reported method that overcomes the difficulty of the extreme resistance of protozoan oocysts to disruption and permits the release of intact meiotic chromosomes. The chromosomes were selected under a light microscope and those selected were stained with phosphotungtic acid and studied by transmission electron microscopy. The authors confirmed 14 chromosomes, which were observed as synaptonemal complexes, and the karyotype was constructed by arranging synaptonemal complexes according to their relative lengths and kinetochore position. Components of the synaptonemal complex, lateral elements, central element, recombination nodules and kinetochore were observed. Measures of the kynetochore, width of the synaptonemal complex, diameter of the recombination nodule and length of the telomeres are given. Minimal and no significant differences were found between measures of chromosomes isolated from different Eimeria tenella strains. To the best of our knowledge, the present investigation for the first time identifies and describes the morphological characteristics of the synaptonemal complex of Eimeria tenella during the meiosis that occurs within the oocysts. In addition, the authors provide evidence of the presence of recombination nodules, suggesting that the recombination process may play an important role in the molecular evolution of this parasite.  相似文献   

5.
Demonstration of the synaptonemal complex for light microscopy has until now been based on staining with silver. After fixation at pH 9-10 it is also possible to visualize synaptonemal complexes with several nonspecific protein stains such as Coomassie brilliant blue, Giemsa, fast green, light green and Stains All. Although staining with silver gives the best contrast between synaptonemal complexes and the background, the other dyes have a number of advantages, such as more even staining, easy extractability, and lower cost than silver.  相似文献   

6.
Demonstration of the synaptonemal complex for light microscopy has until now been based on staining with silver. After fixation at pH 9-10 it is also possible to visualize synaptonemal complexes with several nonspecific protein stains such as Coomassie brilliant blue, Giemsa, fast green, light green and Stains All. Although staining with silver gives the best contrast between synaptonemal complexes and the background, the other dyes have a number of advantages, such as more even staining, easy extractability, and lower cost than silver.  相似文献   

7.
S M Stack  D Roelofs 《Génome》1996,39(4):770-783
Allium porrum L. (cultivated leek) (2n = 4x = 32) is a fertile tetraploid that forms bivalents with pericentric chiasmata at metaphase I. To investigate the basis of this unusual behavior for a tetraploid, we describe the karyotype, axial cores, synaptonemal complexes (SCs), and meiotic nodules of A. porrum. The karyotype appears to be autotetraploid. This conclusion is also supported by presynaptic alignment of axial cores in groups of four and partner trades between pairs of SCs. Numerous early nodules are distributed all along axial cores and SCs during zygonema, but they are lost by late zygonema - early pachynema. Late (recombination) nodules (RNs) are present on SCs near kinetochores throughout the remainder of pachynema. This pattern of RNs corresponds to the pattern of pericentric chiasmata. Pachytene quadrivalents usually are resolved into bivalents because partner trades between SC lateral elements rarely occur between RNs on the same segment of SC. Thus, the patterns of crossing-over and partner trades promote balanced disjunction and high fertility in autotetraploid A. porrum. Rare quadrivalents observed at metaphase I must be due to infrequent partner trades between RNs. Polycomplexes, unusual in their number and size, were observed during zygonema. Key words : synaptonemal complex, recombination nodules, localized chiasmata, polycomplex, Allium porrum.  相似文献   

8.
Two-dimensional spreads of synaptonemal complexes (SCs) from the lower vascular plant Psilotum nudum were examined after staining with uranyl acetate-lead citrate (UP). Staining with UP allows visualization of lateral elements/axial cores (ACs), central elements, kinetochores, and nodules. Numerous darkly stained nodules were associated with forming SCs. In addition to nodules found on the central element of SC segments, other nodules were found at points of convergence between two adjacent ACs. Of these latter nodules, some were obviously associated with a fiber that connected adjacent ACs. No central element material was visible between the ACs, and the nodule complex appeared to be the only structure holding the ACs together. Although the function(s) of nodules during zygotene is unknown, the presence of a nodule-fiber complex that connects adjacent ACs before central element formation suggests that at least some of the nodules may be involved in synaptic initiation.  相似文献   

9.
The use of osmium tetroxide fixation followed by postreatment with p-phenylenediamine gives an opportunity of locating the synaptonemal complex (SC) under the light microscope in mouse testes and Allium cepa anthers. When semi-thin sections from these materials were observed under phase contrast optics or dark field microscopy, fine threads in the pachytene nuclei were clearly visible. Post-staining of semi-thin sections with ammoniacal silver increased the contrast of the SC and allowed for observations using a bright field illumination. Ultrathin sections of osmium tetroxide/ p-phenylenediamine treated material showed that, under the electron microscope, this technique stains preferentially elements of the synaptonemal complex, while the surrounding chromatin remains unstained.  相似文献   

10.
Benjamin C. Lu 《Chromosoma》1993,102(7):464-472
A protocol was developed to spread the synaptonemal complex (SC) of the fungus Neurospora crassa. It involves direct mechanical breakage of meiotic cells before spreading. This technique makes it possible to examine the SC of the same nucleus with both light and electron microscopy. This protocol is potentially applicable for other Pyrenomycetes. The SCs were examined at zygotene, pachytene and diplotene. The central elements and the recombination nodules (RN) were well revealed by silver staining. Ten to 13 RNs were counted at pachytene. The total genomic SC length varied with the stage. This whole mount electron microscopy of the SC is particularly useful for studying chromosomal rearrangements.  相似文献   

11.
Recombination nodules in the oocytes of the chicken, Gallus domesticus   总被引:2,自引:0,他引:2  
Chicken oocytes at pachytene were processed with the microspreading technique (Moses, 1977), and their synaptonemal complex (SC) complements were analyzed by electron microscopy. Ellipsoidal nodules, 140 X 120 nm in diameter, were associated with the central space of synaptonemal complexes. The average number of nodules per pachytene oocyte was 57.5. The number of nodules per bivalent showed a clear linear relationship with SC length, except for the microchromosomes, which showed a single obligatory nodule. The distribution of nodules along the 10 longest SCs was nonrandom, with low frequencies in the vicinity of kinetochores and high frequencies near the telomeres. The microchromosomes showed a single nodule whose average location was 1.21 micron from the kinetochore. In the ZW pair there was a single nodule whose average location was 0.31 micron from the paired telomeres and not more than 0.65 micron from them. The total number of nodules per cell and the number of nodules in each of the five major bivalents showed good agreement with the total number of chiasmata and the number of chiasmata of the major bivalents of roosters. Thus, these nodules share the characteristics of recombination nodules described in other organisms. The single, obligatory, strictly localized recombination nodule found in the pairing end of the ZW pair strongly suggests that recombination between the Z and W chromosomes in the female chicken is a regular process that may be similar to the obligatory recombination between the pairing ends of the human X and Y chromosomes that was recently described in studies using DNA probes.  相似文献   

12.
C. B. Gillies 《Chromosoma》1973,43(2):145-176
Aldehyde fixation followed by staining with phosphotungstic acid produces differential contrast between the synaptonemal complex and the chromatin of maize pachytene bivalents. Centromeres, heterochromatic knobs and large chromomeres are easily recognised. With this and other staining techniques the nucleolus organizer region can be differentiated into two components. — Microsporocyte nuclei at pachytene were serially sectioned and all ten bivalents reconstructed in five nuclei. An idiogram was derived from the mean chromosome (= synaptonemal complex) lengths, the arm ratios, positions of knobs and the nucleolus organizer region. The idiogram agrees well with that published from light microscopic analyses. However, bivalent lengths are only two thirds of those observed by light microscopy of squash preparations. Many telomeres of the bivalents are connected via chromatin to the nuclear envelope, but a varying number of free bivalent ends are observed in all five reconstructed nuclei. — Bivalents heterozygous for inversion 3b were reconstructed. In the presence of abnormal chromosome 10 (K10) the lateral components of the synaptonemal complex of chromosome 3 formed a typical inversion loop, while in one of the nuclei having no K10 the two lateral components of the long arms of chromosome 3 remained unpaired in the region of inversion heterozygosity. The presence of K10, which increases crossing-over frequencies and promotes intimate pairing at the light microscopic level, was thus found to permit formation of complete synaptonemal complexes in the inverted region. The extra terminal portion of the K10 chromosome folded back on itself and formed a morphologically normal synaptonemal complex in this — possibly non-homologously paired — region. The chromatin of centromeres and knobs from different bivalents were sometimes found to fuse, but the synaptonemal complexes transversing the fused centromeres or knobs retained their individuality.  相似文献   

13.
14.
Rat oocytes in the meiotic prophase are studied by means of classical techniques of electron microscopy, preferential staining methods for DNA and RNA and specific enzymatic hydrolysis. The axial cores in leptotene and the lateral arms in the pachytene synaptonemal complex are composed by fibrils that keep a positive contrast after the application of the ethylenediaminetetraacetic acid staining method. They disappear with RNAse treatment, which reveals the presence of chromatin fibrils in the zone occupied by the cores. Preferential staining for DNA corroborates this evidence. Medial arm and lateral-medial fibrils are formed by ribonucleoproteic filaments that form bridges between pairing homologues in the zygotene. In the advanced pachytene stage, the RNA becomes scarce in these structures. No DNA can be detected either in the lateral-medial fibrils or in the medial arm. During diplotene the synaptonemal complex loses its individually and the synaptic space becomes wider and irregular. At the same time, loss of chromatin and a large increase of RNA-containing particles occur. These processes lead to the typical interphasic arrangement of nuclear components seen in the dictyate stage.  相似文献   

15.
鲻鱼早期卵子发生的超微结构研究   总被引:13,自引:0,他引:13  
用电子显微镜技术观察鲻鱼早期卵子发生进入第一次成熟分裂前期联会丝复合体期、粗线期、双线期和网状期卵原细胞和初级卵母细胞生发泡和胸质的超微结构特点。在联会丝复合体期,生发泡内同源染色体配对,联会丝复合体中央出现重组节,胞质中不同发育类型的核仁样及其相关线粒体的分布及其数量可作为划分鲻鱼早期卵子发生各个时期的依据。另外,首次观察到靠近膜细胞有一种不规则形细胞,推测是分泌成熟抑制肽细胞。  相似文献   

16.
A nuclear matrix fraction was prepared from ovaries of the achiasmatic flour moth, Ephestia kuehniella, by removal of the chromatin, using detergent treatment of homogenized ovaries or dissected ovary tips followed by DNase digestion and high salt extraction. Removal of DNA and histones from the nuclei was demonstrated by Feulgen staining and polyacrylamide gel electrophoresis (PAGE), respectively. By light microscopy, ribbon-like structures similar in dimension to the synaptonemal complex were observed in the oocyte after digestion of the chromosomes. Electron microscopic examination of matrix preparations of pachytene cells showed a defined synaptonemal complex structure with both lateral and central elements. Such structures were not found in either the fully differentiated nurse cells or in follicle cells which were exposed to the same preparative technique concurrently. However, in early post-pachytene nurse cells the typical polycomplex structures, formed in these cells from the synaptonemal complex, were found in nuclear matrix preparations. The results suggest an association of synaptonemal complexes with the nuclear matrix.  相似文献   

17.
The synaptonemal complex protein SCP3 is part of the lateral element of the synaptonemal complex, a meiosis-specific protein structure essential for synapsis of homologous chromosomes. We have investigated the fiber-forming properties of SCP3 to elucidate its role in the synaptonemal complex. By synthesis of SCP3 in cultured somatic cells, it has been shown that SCP3 can self-assemble into thick fibers and that this process requires the COOH-terminal coiled coil domain of SCP3, as well as the NH2-terminal nonhelical domain. We have further analyzed the thick SCP3 fibers by transmission electron microscopy and immunoelectron microscopy. We found that the fibers display a transversal striation with a periodicity of ~20 nm and consist of a large number of closely associated, thin fibers, 5–10 nm in diameter. These features suggest that the SCP3 fibers are structurally related to intermediate filaments. It is known that in some species the lateral elements of the synaptonemal complex show a highly ordered striated structure resembling that of the SCP3 fibers. We propose that SCP3 fibers constitute the core of the lateral elements of the synaptonemal complex and function as a molecular framework to which other proteins attach, regulating DNA binding to the chromatid axis, sister chromatid cohesion, synapsis, and recombination.  相似文献   

18.
Electron microscopy of spread maize pachytene synaptonemal complexes   总被引:2,自引:1,他引:1  
C. B. Gillies 《Chromosoma》1981,83(5):575-591
The Counce-Meyer microspreading technique for animal synaptonemal complexes (SCs) has been adapted to allow spreading of the SCs of maize pachytene microsporocytes for examination in the electron microscope (EM). The spread nuclei were well dispersed and flattened, and unstained SCs could be seen with light microscope (LM) phase optics. After PTA or ammoniacal silver staining, the SCs and kinetochores were readily recognized in the EM. Variable degrees of asynapsis, stretching of the SCs, and nonhomologous synapsis of lateral elements were noted, and cases of interlocking of lateral elements or SCs were not uncommon. Distinct lens-shaped thickenings of one or both lateral elements were observed at numerous sites along the SC in most nuclei. — The yield of well spread, complete nuclei, although not high, was sufficient to allow karyotypes to be prepared, based on relative SC lengths and arm ratios. The karyotypes agreed well with published EM and LM determinations, establishing the accuracy of the spreading technique for maize. However, considerable variation in absolute lengths of the SCs was noted. To evaluate the utility of the technique for cytogenetic investigations, two paracentric inversions, and two reciprocal translocations were spread and examined in the EM. The breakpoints estimated from measurements of spread SCs were in agreement with LM determinations.  相似文献   

19.
The mechanism by which homologous chromosomes pair and crossover has been a major unsolved problem in genetics. Thin section electron microscopy of the synaptonemal complex has not provided enough details to allow any significant insight into this problem. Whole mount preparations of the testis of mice, quail, crayfish, and frogs provided a striking improvement in visualization of the morphological features of meiotic chromosomes. These studies, when combined with the use of deoxyribonuclease and trypsin allowed the following conclusions. 1. The synaptonemal complex (lateral and central elements with connecting L-C fibers) is composed of protein. 2. Contrary to common speculation the central element is not the pairing surface of homologous chromosomes. 3. The L-C fibers, averaging 75–100 Å in width, extend from the lateral elements and meet to form the central element which is usually composed of four fibers. 4. During leptotene, homologous axial elements, although unpaired for most of their length, attach next to each other at the nuclear membrane. 5. Short segments of the chromatin fibers attach to the lateral elements. These points of attachment are clustered, producing the chromomeres seen by light microscopy. 6. The chromatin fibers extend out from the lateral element as loops. Lampbrush chromosomes are thus not restricted to oogenesis but are common to all meiotic chromosomes.Since the morphological features of the central element of the synaptonemal complex persist despite extensive deoxyribonuclease digestion, pairing is perhaps best visualized as a two-step process consisting of a) chromosomal pairing during which the proteinaceous synaptonemal complex pulls homologous chromosomes into approximate association with each other, and b) molecular pairing, which probably takes place in the area around the synaptonemal complex.Supported by NIH Grants GM-15886 and C-2568, and The Charles and Henrietta Detoy Research Fellowship.  相似文献   

20.
Synaptonemal complexes analysis was performed using electron microscopy on surface-spread spermatocytes of a bull heterozygous for the 4;8 Robertsonian translocation. In 19 cells examined, the longest autosomal complex showed kinetochores in a central position whereas the remaining autosomal complexes showed terminal kinetochores. Synapsis in the trivalent appeared complete in all cells, and the trivalents usually showed a CIS configuration. The arm ratio varied from 1.05 to 2.04 with an average of 1.32 +/- 0.43. Out of 47 cells showing X-Y bivalents, 34 showed a small synaptonemal complex at one extremity of the X chromosome, and an unstained gap in the Y chromosome. There was no association between the X-Y bivalent and the trivalent. The absence of association would explain the normal spermatogenesis noted in this bull, in contrast to human and mouse carriers of translocations which show impaired spermatogenesis due to the association between the rearranged chromosomes and the sex vesicle. Further studies involving bulls carrying one or more Robertsonian translocations are needed to determine whether this absence of association is a constant feature in cattle.  相似文献   

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