共查询到20条相似文献,搜索用时 15 毫秒
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Reverse transcriptase activity in mature spermatozoa of mouse 总被引:1,自引:0,他引:1
Giordano R Magnano AR Zaccagnini G Pittoggi C Moscufo N Lorenzini R Spadafora C 《The Journal of cell biology》2000,148(6):1107-1113
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电脉冲作用将外源基因导入稀有(鱼句)鲫精子的研究 总被引:5,自引:0,他引:5
将稀有鲫 (Gobiocyprisrarus)精子与重组质粒pCAhLFc线性DNA混合温育 ,经电脉冲处理后与卵子受精 ,孵化出苗。从鱼苗中提取DNA ,经PCR检测 ,2 5 .5 %~ 6 6 .7%鱼苗带有外源基因。在显微镜下观察经电脉冲处理过的精子 ,发现其活力有不同程度下降 ,受精率也有不同程度下降 ,说明不同的电脉冲条件对精子有不同程度的损害作用。精子与外源DNA混合温育 ,经电脉冲处理后 ,用DNA外切酶消化后 ,提取精子DNA ,经PCR检测 ,仍有阳性电泳带 ,证明电脉冲可以促使稀有鲫精子摄入外源基因 相似文献
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Campos VF Komninou ER Urtiaga G de Leon PM Seixas FK Dellagostin OA Deschamps JC Collares T 《Theriogenology》2011,75(8):1476-1481
The objective was to introduce exogenous DNA into commercially sex-sorted bovine sperm using nanopolymer for transfection. In the first experiment, the optimal concentration and ratio of linear-to-circular plasmid was determined for NanoSMGT in unsorted sperm. A second experiment was conducted to transfect exogenous DNA into sex-sorted sperm. Exogenous DNA uptake occurred in a dose-dependent manner (P < 0.05). The optimal amount of DNA was 10 μg/106 cells. The ratios of linear-to-circular plasmid do not influence the uptake by unsorted sperm cells and none of the tested treatments affected sperm motility and viability. Commercially sex-sorted bovine sperm were able to uptake exogenous DNA using nanopolymer; however, both X- and Y-sorted sperm had decreased DNA uptake in comparison to unsorted sperm (P < 0.05). Neither sperm motility nor viability were affected by nanotransfection. In conclusion, nanopolymer efficiently introduced exogenous DNA into commercially sex-sorted bovine sperm; we inferred that these sperm could be used for production of embryos of the desired sex, a technique named NanoSMGT. 相似文献
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The origin and subsequent evolution of new genes have been considered as an important source of genetic and phenotypic diversity in organisms. Dog breeds show great phenotypic diversity for morphological, physiological, and behavioral traits. However, the contributions of newly originated retrogenes, which provide important genetic bases for dog species differentiation and adaptive traits, are largely unknown. Here, we analyzed the dog genome to identify new RNA‐based duplications and comprehensively investigated their origin, evolution, functions in adaptive traits, and gene movement processes. First, we totally identified 3,025 retrocopies including 476 intact retrogenes, 2,518 retropseudogenes, and 31 chimerical retrogenes. Second, selective pressure along with ESTs expression analysis showed that most of the intact retrogenes were significantly under stronger purifying selection and subjected to more functional constraints when compared to retropseudogenes. Furthermore, a large number of retrocopies and chimerical retrogenes that occurred approximately 22 million years ago implied a burst of retrotransposition in the dog genome after the divergence time between dog and its closely related species red fox. Interestingly, GO and pathway analyses showed that new retrogenes had expanded in glutathione biosynthetic/metabolic process which likely provided important genetic basis for dogs' adaptation to scavenge human waste dumps. Finally, consistent with the results in human and mouse, a significant excess of functional retrogenes movement on and off the X chromosome in the dog confirmed a general pattern of gene movement process in mammals which was likely driven by natural selection or sexual antagonism. Together, these results increase our understanding that new retrogenes can reshape the dog genome and provide further exploration of the molecular mechanisms underlying the dogs' adaptive evolution. 相似文献
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Detection of transgene in progeny at different developmental stages following testis-mediated gene transfer 总被引:2,自引:0,他引:2
Yonezawa T Furuhata Y Hirabayashi K Suzuki M Takahashi M Nishihara M 《Molecular reproduction and development》2001,60(2):196-201
We recently reported that exogenous DNA injected into testis as a liposome complex can be transferred into the egg via sperm by natural mating and integrated in the genome (testis-mediated gene transfer: TMGT). Here, we studied the efficiency of each of the several liposomes in associating foreign DNA with sperm, the expression of an introduced gene in early embryos, and the presence of the DNA in fetuses and pups at different ages. The CMV/beta-actin/EGFP fusion gene, encapsulated with different liposomes, was injected into rat testis, and spermatozoa in the cauda epididymis were obtained 1, 4, and 14 days after injection. We tested each of the 8 liposomes, and found that only 2, DMRIE-C and SuperFect, led to the detection of foreign DNA on all of the days examined, with relatively higher ratios of rats having positive sperm. By means of TMGT using either of those two liposomes, more than 80% of morula-stage embryos expressed EGFP, as observed by fluorescence microscopy. Then we detected introduced DNA in the progeny by PCR and Southern dot blot, and found that the ratio of animals carrying the foreign DNA decreased as they developed, and that only a part of postpartum progeny were foreign-DNA-positive with high incidence of mosaicism. These results suggest that, although, the success rate is still limited, foreign DNA could be integrated into the genome of the progeny by TMGT at least under specific experimental conditions, the efficiency of which depends largely on the characteristics of the liposome. The results also suggest that TMGT could be applicable to fetal gene therapy as well as to the generation of transgenic animals. 相似文献
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大球母贝精子介导外源基因转移研究 总被引:9,自引:0,他引:9
将大球母贝(Pinctada maxima Jameson)精子与“全鱼”GH基因重组体pCAgcGH和pCAgcGHc的线性DNA混合,温育30min,经6次、2^7、10kV脉冲电处理后,与卵子受精,得到若干贝苗。从贝苗中提取DNA,经PCR扩增和Southern blot分子杂交表明,部分受体带有外源基因,当与精子温育的外源基因浓度分别为2μg/mL,6μg/mL及18μg/mL时,相应贝苗 相似文献
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Kiyono Sekii Dita B. Vizoso Georg Kuales Katrien De Mulder Peter Ladurner Lukas Sch?rer 《Proceedings. Biological sciences / The Royal Society》2013,280(1757)
Sperm production is a key male reproductive trait and an important parameter in sperm competition theory. Under sperm competition, paternity success is predicted to depend strongly on male allocation to sperm production. Furthermore, because sperm production is inherently costly, individuals should economize in sperm expenditure, and conditional adjustment of the copulation frequency according to their sperm availability may be expected. However, experimental studies showing effects of sperm production on mating behaviour and paternity success have so far been scarce, mainly because sperm production is difficult to manipulate directly in animals. Here, we used phenotypic engineering to manipulate sperm-production rate, by employing dose-dependent RNA interference (RNAi) of a spermatogenesis-specific gene, macbol1, in the free-living flatworm Macrostomum lignano. We demonstrate (i) that our novel dose-dependent RNAi approach allows us to induce high variability in sperm-production rate; (ii) that a reduced sperm-production rate is associated with a decreased copulation frequency, suggesting conditional adjustment of mating behaviour; and (iii) that both sperm production and copulation frequency are important determinants of paternity success in a competitive situation, as predicted by sperm competition theory. Our study clearly documents the potential of phenotypic engineering via dose-dependent RNAi to test quantitative predictions of evolutionary theory. 相似文献
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Structures of four human pseudogenes for U7 small nuclear RNA 总被引:2,自引:0,他引:2
Four U7 RNA-related sequences were isolated from a human genomic DNA library. None of the sequences completely match the published human U7 RNA sequence and all of them contain features typical of reverse-transcribed pseudogenes. 相似文献
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Graziella Iossa Chloris Maury Rachel M. Fletcher Paul E. Eady 《Journal of evolutionary biology》2019,32(7):675-682
In both plants and animals, male gametogenesis is particularly sensitive to heat stress, to the extent that a single hot or cold day can compromise crop productivity or population persistence. In animals, heat stress during development can impact a male's ability to secure copulations and/or his post‐copulatory fertility. Despite such observations, relatively few studies have examined the consequences of developmental temperature on the reproductive behaviour and physiology of individuals. Here, we report for the first time the effects of developmental temperature on the phenotypic expression of both apyrene and eupyrene sperm and the copulatory behaviour of the Indian meal moth, Plodia interpunctella. We show that the length of both apyrene and eupyrene sperm decreases with increasing developmental temperature and that males are less likely to engage in copulation when reared at the highest and lowest temperatures. Where copulation occurred, the duration of copula decreased as male developmental temperature increased. We argue that identification of the mechanisms and consequences of reproductive failure in animals facing heat stress will help understand how wild and domesticated populations will respond to global climate change. We also contend that such studies will help elucidate long‐standing evolutionary questions around the maintenance of genetic variation in traits highly relevant to fitness and the role of phenotypic plasticity in driving the evolution of novel traits. 相似文献
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Intracytoplasmic sperm injection-sperm-mediated gene transfer (ICSI-SMGT) is a useful tool for the production of transgenic mice but is still rather inefficient in farm animals. In the current study, we evaluated the effect of the sperm treatments on the efficiency for producing enhanced green fluorescent protein (EGFP)-expressing pig embryos by ICSI-SMGT. Four different sperm treatments were assayed: (1) fresh (control), (2) frozen-thawing (FT), (3) quick freezing without cryoprotectant agents (QF), and (4) Triton X-100 treatment (TX-100). First, we evaluated the DNA-binding ability and the viability of sperm under the different treatments coincubated with exogenous DNA (EGFP) by flow cytometry. Second, we evaluated the embryo production rate and the efficiency in transgene expression in embryos after using these spermatozoa to fertilize oocytes by ICSI. Sperm treatment significantly increased DNA-binding capacity but reduced sperm viability compared with that of the control group. Treatments damaging the spermatozoa's membranes (QF and TX-100) resulted in a greater capacity of sperm binding exogenous DNA than that after FT treatment (P < 0.01). Similar rates of EGFP-expressing embryos were obtained from the control, FT, and TX-100 groups (37.04 ± 3.52%, 43.54 ± 5.41%, and 29.03 ± 8.29%, respectively), but were significantly higher in the QF group (80.43 ± 5.91%). These results demonstrate that the integrity of the sperm plasma membrane plays a critical role in DNA interaction, and altered plasma membranes facilitate interactions between an injected exogenous DNA and the sperm chromatin. However, severe sperm treatments such as QF and TX-100 may damage the sperm nucleus, induce DNA fragmentation, and/or lead to chromosomal breakage with a detrimental effect on further embryonic development. 相似文献
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Sperm mediated gene transfer in pig: Selection of donor boars and optimization of DNA uptake 总被引:23,自引:0,他引:23
Lavitrano M Forni M Bacci ML Di Stefano C Varzi V Wang H Seren E 《Molecular reproduction and development》2003,64(3):284-291
Transgenic animals are produced primarily by microinjecting exogenous DNA into the male pronuclei of a zygote. Microinjection is successful in mice but not efficient in farm animals, limiting its general utility. We have pursued an alternative technology for producing transgenic animals: Sperm Mediated Gene Transfer (SMGT). Based on our finding that sperm cells bind and internalize exogenous DNA, we used sperm as a vector for transmitting, not only their own DNA, but also, the exogenously-introduced gene of interest to the zygote. SMGT is highly efficient (up to greater than 80%) and relatively inexpensive; it can be used in species refractory to microinjection, whenever reproduction is mediated by gametes. In this report, we describe the procedure for selection of sperm donors and optimization of DNA uptake that are the key steps for the successful outcome of SMGT. We found that the nominal parameters that boar sperm should possess to serve as a good vector for exogenous DNA are the quality of semen based on standard parameters used in conventional animal breeding programs (volume, concentration, presence of abnormal sperm cells, motility at time of collection, and high progressive motility after 2 hr) and the ability of the sperm cells to take up and internalize exogenous DNA. The results described provide significant advances in SMGT technology applied to pigs, so that transgenic pigs can be efficiently obtained. Mol. 相似文献
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In Tribolium flour beetles and other organisms, individuals migrate between heterogeneous environments where they often encounter markedly different nutritional conditions. Under these circumstances, theory suggests that genotype-by-environment interactions (GEI) may be important in facilitating adaptation to new environments and maintaining genetic variation for male traits subject to directional selection. Here, we used a nested half-sib breeding design with Tribolium castaneum to partition the separate and joint effects of male genotype and nutritional environment on phenotypic variation in a comprehensive suite of life-history traits, reproductive performance measures across three sequential sexual selection episodes, and fitness. When male genotypes were tested across three nutritional environments, considerable phenotypic plasticity was found for male mating and insemination success, longevity and traits related to larval development. Our results also revealed significant additive genetic variation for male mating rate, sperm offence ability (P(2)), longevity and total fitness and for several traits reflecting both larval and adult resource use. In addition, we found evidence supporting GEI for sperm defence ability (P(1)), adult longevity and larval development; thus, no single male genotype outperforms others in every nutritional environment. These results provide insight into the potential roles of phenotypic plasticity and GEI in facilitating Tribolium adaptation to new environments in ecological and evolutionary time. 相似文献
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Willian T. A. F. Silva Paula Sez‐Espinosa Stphanie Torijo‐Boix Alejandro Romero Caroline Devaux Mathilde Durieux María Jos Gmez‐Torres Simone Immler 《Journal of evolutionary biology》2019,32(6):535-544
Sperm function and quality are primary determinants of male reproductive performance and hence fitness. The presence of rival males has been shown to affect ejaculate and sperm traits in a wide range of taxa. However, male physiological conditions may not only affect sperm phenotypic traits but also their genetic and epigenetic signatures, affecting the fitness of the resulting offspring. We investigated the effects of male‐male competition on sperm quality using TUNEL assays and geometric morphometrics in the zebrafish, Danio rerio. We found that the sperm produced by males exposed to high male–male competition had smaller heads but larger midpiece and flagellum than sperm produced by males under low competition. Head and flagella also appeared less sensitive to the osmotic stress induced by activation with water. In addition, more sperm showed signals of DNA damage in ejaculates of males under high competition. These findings suggest that the presence of a rival male may have positive effects on sperm phenotypic traits but negative effects on sperm DNA integrity. Overall, males facing the presence of rival males may produce faster swimming and more competitive sperm but this may come at a cost for the next generation. 相似文献
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Jessica H. Hadlow Jonathan P. Evans Rowan A. Lymbery 《Journal of evolutionary biology》2020,33(6):797-807
Sperm cells exhibit extraordinary phenotypic diversity and rapid rates of evolution, yet the adaptive value of most sperm traits remains equivocal. Recent findings suggest that to understand how selection targets ejaculates, we must recognize that female‐imposed physiological conditions often alter sperm phenotypes. These phenotypic changes may influence the relationships among sperm traits and their association with fitness. Here, we show that chemical substances released by eggs (known to modify sperm physiology and behaviour) alter patterns of selection on a suite of sperm traits in the mussel Mytilus galloprovincialis. We use multivariate selection analyses to characterize linear and nonlinear selection acting on sperm traits in (a) seawater alone and (b) seawater containing egg‐derived chemicals (egg water). Our analyses revealed that nonlinear selection on canonical axes of multiple traits (notably sperm velocity, sperm linearity and percentage of motile sperm) was the most important form of selection overall, but importantly these patterns were only evident when sperm phenotypes were measured in egg water. These findings reveal the subtle way that females can alter patterns of selection, with the implication that overlooking environmentally moderated changes to sperm, may result in erroneous interpretations of how selection targets phenotypic (co)variation in sperm traits. 相似文献