首页 | 本学科首页   官方微博 | 高级检索  
相似文献
 共查询到20条相似文献，搜索用时 750 毫秒
1.
Large yellow croaker, Pseudosciaena crocea, exhibit sexually dimorphic growth, with females growing faster and reaching larger adult sizes than males. Thus, development of techniques for preferentially producing females is necessary to optimize production of these species. We have established a protocol to produce all-female croaker P. crocea through induction of meiotic gynogenesis with homologous sperm. The first set of experiments investigated the ultra-violet (UV) irradiation on sperm motility and duration of sperm activity to determine the optimal UV dosage for genetic inactivation of sperm, yet retaining adequate motility for activation of eggs. Milt from several males was diluted 1:100 with Ringer’s solution and UV irradiated with doses ranging from 0–150 J cm−2. The results indicated that motility and duration of activity generally decreased with increased UV doses. At UV doses greater than 105 J cm−2, after fertilization, motility was <10% and fertilization rates were significantly lower. Highest hatching rate was obtained at 75 J cm−2. A second set of experiments was carried out to determine appropriate conditions of cold shock for retention of the 2nd polar body in P. crocea eggs after fertilization with UV-inactivated sperm by altering the timing, temperature and duration of shock. At 20°C, shock applied at 3 min after fertilization resulted in higher survival rate of larvae at 6 h after hatching. Results of different combinations of three shock temperatures (2°C, 3°C or 4°C) and five shock durations (4 min, 8 min, 12 min, 16 min or 20 min) at 3 min after fertilization demonstrated that shocks of 12 min gave highest production of diploid gynogens. Statistical analysis revealed that maximum production of diploid gynogens (44.55 ± 2.99%) were obtained at 3°C. The results of this study indicate that the use of UV-irradiated homologous sperm for activation of P. crocea eggs and cold shock for polar body retention is an effective method for producing gynogenetic offspring.  相似文献   

2.
In the present study, methodology of gynogenetic induction in spotted halibut were developed and optimized; the sex ratio of putative meiotic gynogenetic diploids was determined using AFLP-based molecular sexing technique; the homozygosity of gynogenetic population was assessed as opposed to cultivated population. The results showed that high percentage of meiotic gynogenetic diploids were generated when the eggs fertilized with irradiated heterologous sea perch frozen sperm (30–50 mJ cm−2) were cold shocked in sea water of −1°C for 40–75 min at 5 min after fertilization. About 15,200 diploid gynogenetic larvae were achieved and they exhibited normal morphology similar to diploid control. The gynogenetic diploids were 100% female, which first confirmed the female homogamete (XX/XY) sex determination in spotted halibut. The genetic analysis showed that the average H O was, respectively, 0.404 and 0.724 in gynogenetic population and cultivated population, indicating an increase of homozygosity in gynogenetic population.  相似文献   

3.
  相似文献   

4.
The cytological process of induced gynogenetic development and subsequent chromosome duplication by a cold shock treatment was observed in Japanese flounder Paralichthys olivaceus (Temminck et Schlegel). Mature eggs were at the metaphase of the second meiosis when inseminated with ultraviolet (UV)-irradiated sperm of red sea bream Pagrus major . After the beginning of cold shock treatment, the previously visible spindle became invisible, probably due to the side effect caused by cold shock treatment. The chromosomes at the centre of the metaphase plate were condensed. This condition continued during the duration of the cold shock treatment and several minutes after it. The release of the second polar body was blocked and it developed into a female-like pronucleus. Then, it fused with the female pronucleus to generate a diploid zygotic nucleus, and the egg exhibited the first mitosis. Consequently, the haploid female chromosome set of the egg was doubled by the inhibition of the second polar body release. There was a significant delay in developmental time in the gynogenetic eggs when compared with that in the normal eggs. From the time of insemination to early cleavage, the UV-irradiated heterospecific sperm nucleus remained condensed.  相似文献   

5.
Triploid African catfish (Clarias gariepinus) produced through cold shock 3 min post fertilization were compared to diploid offspring of the same parentage at 66 days of age. Triploid fish were significantly shorter (11%) and weighed less (18%) than diploids but showed no significant difference in mortality or cannibalism, which can be an important source of losses under aquaculture conditions. Erythrocytes of triploid fish were more oval shaped than the normal spherical shape of diploid erythrocytes, were significantly larger (38%) and had nuclei that were significantly larger (25%) than observed in diploid fish. Erythrocyte morphological characteristics show potential as an indicator for the detection of triploidy in African catfish.  相似文献   

6.
This study reports the results on induced meiotic diploid gynogenesis and female homogametic nature in the Indian catfish, Heteropneustes fossilis. The eggs of H. fossilis were inseminated with conspecific sperm. The sperm suspension was diluted to 1 × 107 sperm mL−1 in Hanks balanced salt solution. Sperm were irradiated under UV light, with the exposure time ranging from 15 to 360 s (7500 ergs mm−2 for 60 s). The genetic inactivation of paternal chromosomes was confirmed by chromosome counting from the larval cells and the larvae also had a characteristic haploid syndrome. A typical ‘Hertwig effect’ in the yield of hatched larvae was observed with doses of UV exposure >75 s (9375 ergs mm2). Larvae resulting from sperm UV irradiated above 120 s (15 000 ergs mm2) were 100% haploids. Application of heat shock to the activated eggs was effective in suppressing the release of the second polar body (meiotic gynogenesis) and resulted in diploid gynogenetic larvae morphologically identical to those of the control. The best yield of diploid gynogens (49.3% with respect to the control) was found to be at 6 min after egg activation and the heat shock at 41 °C for a 1-min duration, at an ambient water temperature of 27 °C. A total of 113 diploid gynogenetic fry from seven different female fish were reared and subjected to sexing. All gynogenetic fish were female in contrast to the control, which had a mean sex ratio of 56.7% females (which was not significantly different from 50% female). From these results, the sex determination mechanism in H. fossilis was presumed to be female homogamety.  相似文献   

7.
Mass production of fast‐growing, all‐female muskellunge Esox rnasquinongy by gynogenesis requires optimized techniques of preventing second polar body extrusion. Heat, cold, and pressure shocks were evaluated for their efficiency of doubling the maternal genome. Muskellunge eggs (20–40 g) were activated with 1 mL ultraviolet (UV)‐irradiated (1,248 J/ m2) heterologous sperm of yellow perch Perca flavescens. Survival and ploidy (by flow cytometry) were determined during the eyed‐stage. Cold shocks of 1.3 × 1 C were applied at 5 or 20 min after gamete activation with water (time of initiation, TI) for a duration of 150 min and pressure shocks of 48,263 or 55,158 kPa (7,000 or 8,000 psi, respectively) at a TI of 4 rnin for 12 min. These shock treatments resulted in 43.7–95.0% diploid gynogens with corresponding yield of diploid gynogen (percent diploid gynogens × total percent survival) of 2.6–11.I%. Cold shocks applied at TI of 5 or 20 min after activation resulted in statistically similar percent survival, percent diploid gynogens, and yield of diploid gynogens. Heat shocks of 31 × 0.1C applied at a TI of 5 to 15 min for a duration of 5 min resulted in 4.8–21.1% diploid gynogens with yields of 0.1–0.4%. Cold and pressure shocks have better potential than heat shock for preventing the second polar body extrusion. Muskellunge eggs activated with UV‐irradiated yellow perch sperm, but not exposed to shock, resulted in 100% haploids with survival of 2.3–5.8%. No viable embryos were produced from the hybrid cross between muskellunge and yellow perch, thus, all diploids produced after the shock treatments were unambiguous meiotic gynogens. Muskellunge eggs fertilized with fresh muskellunge sperm (controls) showed 60.4–64.0% survival to the eyedstage and 100% diploidy. Considering that the sex‐determining mechanism in muskellunge follows the WZ female, ZZ male system, future efforts should be directed to test the efficiency of cold and pressure shocks for mass‐producing gynogenetic super female (WW) muskellunge.  相似文献   

8.
The aim of this study was a comparison of key haematological features of diploid (2n) and triploid (3n) Caspian salmon (Salmo trutta caspius). Morphometric indices of erythrocytes were determined on blood smears by light microscopy. Triploidy significantly (P < 0.001) increased all morphometric indices measured in the erythrocytes including cell size, cell surface area, and cell volume. The increase in cell size was larger for the major (27%) axis than for the minor (22%) axis, thus making erythrocytes of 3n Caspian salmon more ellipsoidal. The estimated increase in erythrocyte nuclear volume (87%) was bigger than the theoretical expected 50% increase. Haematological indices were measured manually by hemocytometry. Triploids had lower numbers of red blood cells (RBC: 1,120,000 cells/mL in 2n vs. 700,000 cells/mL in 3n; P < 0.001) but they were larger in size (mean erythrocytic volume [MEV]: 363.1 nm3 in 2n vs. 483.3 nm3 in 3n; P < 0.001). The decrease in RBC number was not compensated by the increase in MEV and, thus, triploidy affected the haematocrit (Hct: 38.8% in 2n vs. 33.06% in 3n; P < 0.05). Total blood hemoglobin concentration was lower in triploid fish (Hb: 9.9 g/dL in 2n vs. 8.9 g/dL in 3n; P < 0.05). In contrast, mean erythrocytic hemoglobin (MEH: 95 μg in 2n vs. 133.2 μg in 3n; P < 0.001) was higher for 3n Caspian salmon as a result of their larger erythrocytes, although MEH concentration (MEHC: 0.26 g/dL in 2n vs. 0.27 g/dL in 3n) did not significantly differ (P > 0.05). White blood cell (WBC) counts (lymphocytes and neutrophiles) were measured and WBC/RBC ratios were calculated. There were no significant differences in WBC (15,710 cells/mL in 2n vs. 12,683 cells/mL in 3n; P > 0.05), lymphocytes, and neutrophils as %WBC as well as WBC/RBC ratios between two ploidy levels (P > 0.05). Triploid Caspian salmon showed higher erythrocyte abnormalities such as ‘twisted’, ‘tailed’, and ‘anucleated’ cells as well as high portions of immature RBC in blood smears in comparison with diploids (P < 0.001).  相似文献   

9.
From the two species of bivalves, Calyptogena soyoae around a cold seep and Bathymodiolus septemdierum near hydrothermal vents in the sea, sterols were isolated using high-pressure liquid chromatography analysis of the lipid fraction guided by characteristic 1H NMR signals of the sterol skeleton. The minor sterol composition of C. soyoae included 24-methylenecycloartanal, cycloeucalenol, and obutusifoliol, which are known phytosterols. From B. Septemdierum, lathosterol and cholesterol as main sterols together with more diverse sterols were obtained. The difference between these species and their sterol contents is most likely because of feeding modes and metabolism of nutrients from their habitat.  相似文献   

10.
The effect of thermal history (16 and 20°C) on growth of juvenile turbot, Scophthalmus maximus (initial mean weight 72.6 g, n = 157) was studied. Fish were divided into four groups, two groups remaining at constant temperature (C16, C20), while fish in the other groups were transferred from either 16 to 20°C (F16-20) or from 20 to 16°C (F20-16). Between 35 and 42 fish in each tank were individually tagged at the start of the experiment. The final mean weights were significantly higher in the F20-16 group (230 g) than in the C20 (213 g), F16-20 (211 g) and C16 (205 g) groups. The overall growth rate was highest in the F20-16 group (1.17% day−1) but comparable in the three other groups (1.00–1.04% day−1). Our findings indicate that, even at near-optimal temperature for a given size, the temperature history of the fish may influence future growth. Based on these indications, we conclude that as turbot grow larger, the temperature should be reduced to take advantage of the change in optimal temperature for growth with increasing fish size rather than rearing at constant temperatures.  相似文献   

11.
Quantifying spermatozoa concentration in milt samples is very important for aquaculture purposes, particularly when equal numbers of gametes are needed for experimental purposes, and different methods can be used. Haemocytometer and Coulter counting are the most precise methods to count spermatozoa, but they are very difficult to use when large numbers of samples are involved. A standard curve of spectrophotometric absorbance (A) against sperm concentration ([S]) was used herein instead, and an equation for estimating spermatozoa concentration for the mussel Mytilus edulis L. was obtained. The best wavelength to measure mussel spermatozoa was 320 nm, with a precision two or three times higher than at other wavelengths. The equation for spermatozoa concentration was A = 8.592 × 10−5 [S] + 0.0190 (r 2 = 0.9979). Recommendations for choosing the best wavelength are discussed.  相似文献   

12.
An investigation was conducted to evaluate the annual variation in potential predation impact (PPI) on larval and juvenile Pseudopleuronectes yokohamae by Crangon uritai in Hakodate Bay using predator-prey size relationships. Laboratory experiments were designed to estimate the favorable prey size of C. uritai through back-calculation of body length (BL) of P. yokohamae from sagittal otolith diameter observed in the stomachs of C. uritai. The most favorable prey-predator size ratio (BL of P. yokohamae-total length (TL) of C. uritai) class was 0.15–0.19, and ranged from 0.12–0.31. There was a significant positive correlation between the BL increase of P. yokohamae and the bottom water temperature in the field, such that BLs stagnated during the cold year of 1999 from April to June, and increased during the warm year of 2002. In contrast, no significant correlation was found between the TL increase of C. uritai and the bottom water temperature. Moreover, there were no significant differences in the mean TL of C. uritai between months (April–June) or years (1998–2002). Variation of PPI depended on the relationships between the growth rates of P. yokohamae and bottom water temperature. Therefore, the warm year of 2002 resulted in higher potential predation impact (PPI≥20), and it was at least 20 days shorter than that of the cold year of 1999. These results suggest that increased bottom water temperature in the nursery area was one of the most important factors for cumulative predation loss.  相似文献   

13.
The changes in the biochemical compositions and enzymatic activities of rotifer (Brachionus plicatilis) and Artemia, enriched and stored at 4°C temperature, were determined. The total starvation period was 16 h and samples were taken at the end of the 8th and 16th hours. In present study, the rotifer and nauplii catabolized a large proportion of the protein during the enrichment period. Lipid contents of both live preys increased during the enrichment period and decreased in nauplii and metanauplii throughout the starvation period but lipid content of the rotifer remained relatively constant during the starvation period. The changes observed in the amino acid compositions of Artemia and the rotifer were statistically significant (P < 0.05). The conspicuous decline the essential amino acid (EAA) and nonessential amino acid (NEAA) content of the rotifer was observed during the enrichment period. However, the essential amino acid (EAA) and nonessential amino acid (NEAA) contents of Artemia nauplii increased during the enrichment period. The unenriched and enriched rotifers contained more monounsaturated fatty acid (MUFAs) than polyunsaturated fatty acid (PUFAs) and saturated fatty acids (SFA). However, Artemia contained more PUFAs than MUFAs and SFA during the experimental period. A sharp increase in the amounts of docosahexaenoic acid (DHA) during the enrichment of the rotifer and Artemia nauplii was observed. However, the amount of DHA throughout the starvation period decreased in Artemia metanauplii but not in Artemia nauplii. Significant differences in tryptic, leucine aminopeptidase N (LAP), and alkaline phosphatase (AP) enzyme activities of Artemia and rotifer were observed during the enrichment and starvation period (P < 0.05). The digestive enzymes derived from live food to fish larvae provided the highest contribution at the end of the enrichment period. In conclusion, the results of the study provide important contributions to determine the most suitable live food offering time for marine fish larvae. Rotifer should be offered to fish larvae at the end of the enrichment period, Artemia nauplii just after hatching and before being stored at 4°C, and Artemia metanauplii at the end of the enrichment and throughout the starvation period.  相似文献   

14.
The present study investigated the effects of sequential collection of milt, time of post-mortem storage and anesthesia on rainbow trout (Oncorhynchus mykiss) sperm motility parameters (using computer-assisted sperm analysis – CASA) as well as seminal plasma osmolality and sperm concentration. The post-mortem storage and time of anesthesia altered motility characteristics of rainbow trout sperm to different extents. The moderate impact of time of anesthesia was manifested in a shortened duration of sperm motility after 10 min exposure of fish to anesthetic. The prolonged post-mortem storage (≥40–60 min), in addition to lowering sperm motility duration, also significantly influenced sperm motility parameters, such as sperm velocities, percentage of motile sperm and sperm trajectory parameters. These results clearly demonstrate that when milt from sacrificed fish is used for sperm motility studies, the time of post-mortem storage significantly alters sperm motility characteristics. Since sperm motility rate and swimming velocity could predict fertilizing ability, detrimental effects of prolonged post-mortem storage may lead to reduced fertilization success. Sperm concentration and seminal plasma osmolality were lower in the first fractions and increased with successive collections of milt. It suggests the presence of urine contamination of the first milt fractions which were collected by stripping. Therefore, testing of sperm concentration and/or seminal plasma osmolality should be mandatory while handling stored milt.  相似文献   

15.
We have examined the effects of Tribulus terrestris (TT), a non-toxic herb, on sex reversal in the Convict Cichlid Cichlasoma nigrofasciatum with the aim of introducing a new environmentally friendly method for masculinization in C. nigrofasciatum. TT is a natural plant product that elevates the testosterone levels in humans and animals. Different concentrations (0.0, 0.10, 0.20 and 0.30 g/l) of TT extract were tested for their effect on sex reversal in C. nigrofasciatum by immersing newly hatched offspring once weekly for 2 months in TT extract. Of the dosages used in the present study, 0.30 g/l TT was the most effective in terms of masculinization, resulting in a maximum male ratio of 87.23% (P < 0.001). Sex ratios of 79 and 85% at 0.10 and 0.20 g/l TT, respectively, were also significantly different from the expected 1:1 ratio (P < 0.001). Histological examinations revealed that the testes of fish treated with TT extract contained all stages of spermatogenesis, clearly demonstrating that the administration of TT extract to C. nigrofasciatum stimulated spermatogenesis. Total survival rates in all treatments and the control were uniformly high, ranging from 88.57 to 90% (P > 0.05). We concluded that TT had no negative effect on the survival rate of C. nigrofasciatum. In addition, all groups of TT-treated fish exhibited successful growth acceleration compared to the control group, but only the 0.30 g/l TT treatment significantly improved the growth rate of C. nigrofasciatum. (P < 0.01). Sex reversal in C. nigrofasciatum demonstrated that TT-treated 0-day-old larvae showed successful sex reversal, spermatogenesis and a better growth rate than untreated progenies.  相似文献   

16.
Compensatory growth following stress is a strategy aquatic animals use to adjust themselves to a variable environment. Studies on the recovery growth of aquatic animals are not only of theoretical value in ecophysiology and evolution, but also important to applications in aquaculture and fisheries resource management. In this experiment, juvenile Fenneropenaeus chinensis with an initial average body weight of about 3.72 g were exposed to hypoxic water (about 2.08 and 3.11 mg/l of dissolved oxygen (DO) content) for 10 days and then switched to normoxic water (about 5.63 and 5.59 mg/l DO). Compared with juveniles in normoxia, juveniles in the hypoxia period allocated a greater proportion of energy to metabolism and exuviations but allocated less energy to daily metabolism per gram shrimp weight (J/g/day). This reduced feed conversion efficiency and feeding rate. Finally, F. chinensis suffered growth depression. The juveniles completely compensated for hypoxia-induced growth depression in 30 days after being switched into normoxic water and the compensation was achieved mainly by hyperphagia and slightly by improvement of feed conversion efficiency. During the recovery period the hypoxic-stressed shrimp showed higher daily metabolic energy (J/g/day) than controls (P < 0.05). Which means the stressed shrimp had more energy for feeding-related activities. So hyperphagia was observed. Energy analysis indicated that F. chinensis improved feed-conversion efficiency mainly by reducing the percentage of energy lost in feces and exuviations. The results showed that short-term non-lethal hypoxia would not affect the growth of juvenile F. chinensis if there were enough time for the stressed shrimp to recover. This suggested F. chinensis was capable of adapting to DO fluctuation to some extent and short-term non-lethal hypoxia would not have an obvious effect on natural, released, and cultured shrimp stock.  相似文献   

17.
Effect of temperature on the development of eggs of round herring Etrumeus teres was experimentally examined to construct a temperature-dependent egg development model. Mature fish were collected in the field and their eggs were artificially fertilized onboard. The eggs were incubated at nine temperatures set between 14.0 and 25.0°C. All eggs at the lowest three temperatures, 14.0°C, 15.0°C, and 16.0°C, ceased development and died at various stages before hatching. Durations required to hatching after fertilization ranged from 38.0 h at 25.0°C to 90.0 h at 17.5°C. The temperature-dependent egg development model, i.e., egg age in hours (y i,t ) at the ith stage and temperature t (°C), was expressed as: y i,t  = 4.604 × exp(−0.100 × t −0.129 × i) × i 2.593. From the application of the model to early-stage eggs collected in the field, it is concluded that round herring starts spawning immediately after sunset and almost completes spawning by midnight. The temperature-dependent egg development model and the daily pattern of spawning presented in this study are essential tools for developing the daily egg production method to estimate the spawning stock biomass.  相似文献   

18.
The blue mussel Mytilus edulis is a commercially important species whose fishery and culture generally relies on natural spat collection. Hatchery-production could provide an alternative source of seed, enabling reliable expansion of the industry. Mussel spawning and larval rearing trials were carried out to optimise elements of hatchery production. Culturing fertilised eggs at low density (20–200 eggs cm−2) rather than high density (400–720 eggs cm−2) significantly improved the quality of first veliger larvae and differences in this improvement were evident between the eggs from different females (maternal effects). Veliger larval growth at 17 or 21°C was significantly faster than growth at 14°C. Feeding veliger larvae an identical total ration either daily or at 2–3 day intervals did not significantly affect their growth. Different microalgal diets (1: Isochrysis sp. (clone T-ISO), 2: Chaetoceros calcitrans forma pumilus, 3: C. muelleri, 4: mixed Isochrysis sp. (clone T-ISO) and C. calcitrans f. pumilus, and 5: mixed Isochrysis sp. (clone T-ISO) and C. muelleri) were tested on veliger larval growth and mixed diets outperformed single-species diets.  相似文献   

19.
Artificial and natural hybridization of dojo loach Misgurnus anguillicaudatus (2N = 50, DD for short) and large-scale loach Paramisgurnus dabryanus (2N = 48, PP for short) are well-grown. However, these hybrid loaches have not yet been examined for fertility and ploidy of gametes. Here, histological observations, artificial propagation, observations of embryonic development, larval morphology, and ploidy analyses were conducted to determine the fertility and ploidy of gametes of allodiploid (DP for short) and allotriploid (DDP for short) loaches, produced by DD females × PP males and induced from fertilized eggs of DD females × PP males by cold shock to prevent the second polar body release, respectively. The ovaries of DP and DDP included smaller number of eggs when compared with those of the control DD, while full-grown oocytes were observed. Testes of these two loaches were delayed-developed without spermatids or mature spermatozoa. Results obtained here showed that DP and DDP were fertility-weakened female and sterile male. Moreover, DP females and DDP females could, respectively, produce few viable haploid eggs and few viable haploid and diploid eggs. This study will provide valuable information for fish hybrid researches.  相似文献   

20.
Effects of the starting time and duration of cold shock, as well as the source of heterogenic sperm on the percentage of viable gynogenic larvae (PVGL) in tench were studied. The DNA in sperm of red common carp (Cyprinus carpio var singuonensis) was inactivated by ultraviolet radiation prior to use to induce meiotic gynogenetic development in tench. In experiment 1, tench eggs were cold-shocked for 30 min starting at 1, 3, 5, 7, 10 and 15 min post activation. In experiment 2, cold shock began 5 min after activation and lasted for 10, 20, 30, 40, 50 min, respectively. Each experiment was run in triplicate using 3 tench females, and one group not treated with cold shock was included in each experiment to serve as a control group. In experiment 3, sperm of bigmouth buffalo (Ictiobus cyprinellus), red common carp or grass carp (Ctenopharyngodon idella) was used to induce gynogenesis in tench with a 20-min cold shock starting 5 min post activation. The results showed that PVGL from the control group was very low (0.11–0.47%). In experiment 1, the highest average PVGL (9.60%) was observed when cold shock treatment was applied 5 min post activation. When cold shock treatment was started 5 min post activation, duration of cold shock affected PVGL. Cold shock lasting 20 min resulted in the highest average PVGL (12. 57%) among the selected duration of cold shock studied in experiment 2. The average PVGL was 2.3, 8.6, and 9.3%, respectively, for eggs induced by sperm of bigmouth buffalo, red common carp and grass carp. Average PVGL was significantly lower for eggs induced by sperm of bigmouth buffalo, compared with that for eggs induced by sperm of the other two species. However, average PVGL were similar for eggs induced by sperm of red common carp and grass carp. In summary, the optimal conditions for gynogenesis in tench include the use of irradiated sperm of grass carp to activate the eggs and cold shock of 20 min starting 5 min post activation. Since female tench grow much faster to a larger size than male tench, gynogenesis of tench holds great potential for production enhancement.  相似文献   

设为首页 | 免责声明 | 关于勤云 | 加入收藏

Copyright©北京勤云科技发展有限公司  京ICP备09084417号