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 36 毫秒
1.
We sequenced 540 nucleotides of the last exon in the ZFY/ZFX gene in two males and two females for eight cetacean species; four odontocetes (toothed whales) and four mysticetes (baleen whales). Based upon the obtained nucleotide sequences, we designed two sets of oligonucleotide primers for specific amplification of the ZFX and the ZFY sequence in odontocetes and mysticetes, respectively. Each primer set consisted of three oligonucleotides; one forward-orientated primer, which anneals to the ZFY as well as the ZFX sequence, and two reverse-orientated primers that anneal to either the ZFX or the ZFY sequence. The resulting two amplification products (specific for the ZFY and ZFX sequences) can be distinguished by gel-electrophoresis through 2% NuSieve™. The accuracy of the technique was tested by determination of gender in 214 individuals of known sex. Finally we applied the technique to determine the sex of 3570 cetacean specimens; 2284 humpback whales, 315 fin whales, 37 blue whales, 7 minke whales, as well as 592 belugas, 335 narwhals and 25 harbour porpoises.  相似文献   

2.
3.
An inexpensive, time-saving and reliable method, polymerase chain reaction with confronting two-pair primers (PCR-CTPP), was developed for sex identification in tiger (Panthera tigris) based on zinc finger alleles (ZFX/ZFY). A site of “C/G” transversion representing fixed differences that discriminated between ZFX and ZFY exons among felids was identified for primers designing. This primer set was successfully tested on samples including blood, shed hairs, dried skin, and stool which contained potential contamination caused by prey DNA. Cross species tests shown that this primer set was also useful for sex identification in four other endangered felids.  相似文献   

4.
Qiao Y  Zou F  Wei K  Yue B 《Zoological science》2007,24(5):493-495
We describe a rapid sex-identification method for the forest musk deer (Moschus berezovskii) using PCR based on zinc-finger protein-encoding genes (ZFX/ZFY) located on the X and Y chromosomes. Fragments of the ZFX and ZFY genes were amplified and sequenced. The ZFX and ZFY fragments were identical in length and 94% similar in nucleotide sequence. Specific primers for forest musk deer sex identification were designed on the basis of sequence differences between ZFX and ZFY. All the primers were multiplexed in single-tube PCR. Both male and female forest musk deer showed amplification bands of 447 bp and 212 bp separated in agarose gels. A sex-specific 278-bp band was amplified only from males. These results show that testing by PCR for the presence of the 278-bp sequence is a rapid and reliable method for sex identification.  相似文献   

5.
Sequence polymorphism of homologues ZFX and ZFY, in a 604-base pair exon region, was examined in 10 known males and 10 known females across seven cetacean families and used to design a simple, highly sensitive and widely applicable fluorescent 5' exonuclease assay for gender determination in cetaceans. Multiplex amplification, cloning, and sequencing of these previously uncharacterized regions revealed (i) eight fixed differences between ZFX and ZFY, (ii) 29 variable sites between ZFX and ZFY and (iii) very low interspecific nucleotide diversity for both ZFX and ZFY across all families examined. We developed a 5' exonuclease assay that produces a small (105 bp) polymerase chain reaction (PCR) product from both the X and the Y chromosome orthologs, and used double-labelled fluorescent probes to distinguish between the two genes in a real-time PCR assay that is highly reproducible and sensitive. We demonstrated sex specificity for 33 cetacean species in nine families. Given the availability of conserved primers and sequence information for many mammalian species, this approach to designing sexing assays for a wide range of species is both practical and efficient.  相似文献   

6.
毛冠鹿ZFY、ZFX基因片段的克隆与性别鉴定   总被引:7,自引:0,他引:7  
蒋华云  曹祥荣  张锡然  胡均  徐春茂 《遗传》2004,26(4):465-468
根据人和鼠性别分化相关的ZFY、ZFX基因序列设计引物,以雌雄毛冠鹿的基因组DNA为模板进行PCR扩增,将扩增产物克隆到pMD18T上,获得ZFY、ZFX重组克隆,并测定了ZFY、ZFX基因片段的序列,序列比较显示两者同源性达 91%,仅在少数位点有差异,以此确定AvaⅡ为ZFX上特异酶切位点,通过PCR扩增和AvaⅡ特异酶切对毛冠鹿性别进行鉴定。 Abstract: According to the human sex differentiation related ZFY and ZFX genes, a pair of primers were designed , and fragments were amplified from the genomic DNA of male or female tufted deer. Subsequently the amplified fragments were cloned into the vector pMD18T and were sequenced. It is found that the sequences of ZFY gene and ZFX gene have 91% homology. Based on the different nucleotides, restriction site of AvaⅡ was found to be specific to ZFX gene. The results show that the combination of PCR with AvaⅡ digestion is a simple and sensitive way to identify the tufted deer sex.  相似文献   

7.
A growing body of evidence suggests the involvement of sex chromosome genes in mammalian development. We report the cloning and characterization of the complete coding regions of the bovine Y chromosome ZFY and X chromosome ZFX genes, and partial coding regions of porcine and equine ZFX and ZFY genes. Bovine ZFY and ZFX are highly similar to each other and to ZFX and ZFY from other species. While bovine and human ZFY proteins are both 801 amino acids long, bovine ZFX is 5 amino acids shorter than human ZFX. Like in humans, both bovine ZFY and ZFX contain 13 zinc finger motifs and belong to the Krueppel family of C2H2-type zinc finger proteins. The internal exon-intron organization of the bovine, porcine and equine ZFX and ZFY genes has been determined and compared. Within this region, the exon lengths and the positions of the splice sites are conserved, further suggesting a high evolutionary conservation of the ZFX and ZFY genes. Additionally, new alternatively spliced forms of human ZFX have been identified.  相似文献   

8.
Schwerin M  Pitra C 《Theriogenology》1994,41(2):553-559
We describe a quick and efficient method of determining the sex of DNA samples in the hyena. By choosing primers from sequences that are conserved between the human and bovine ZFY and ZFX genes, we amplified a 448 bp fragment from 1 male and 2 female hyenas. Using comparative sequencing, single base pair polymorphisms between the amplified ZFY and ZFX were established. Restriction fragment length polymorphism (RFLP) analysis with PstI and TaqI confirmed the sequence data and yielded specific banding patterns between the 2 sexes in the hyena.  相似文献   

9.
Buffalo Y-chromosome specific repetitive DNA (BuRY.I) was cloned and sequenced in order to develop a sensitive method for sexing of buffalo preimplantation stage embryos using polymerase chain reaction (PCR). A highly sensitive and reliable sex determination assay using a primary (BRY.I), nested (BuRYN.I) and multiplex (BuRYN.I, ZFX/ZFY) PCR was developed. The BRY.I and BuRYN.I primers are targeted to amplify Y-specific sequences, while the ZFX/ZFY loci was amplified to serve as a positive control for both male and female samples. Accuracy of the sex determination assay was initially verified with genomic DNA obtained from blood of known gender. Further sensitivity and reproducibility of the assay was examined using DNA obtained from 1 or 2 blastomeres to demi embryos. Altogether, 80 IVF-derived embryos ranging from the 2 to 4 cell to the blastocyst stage were used for sex determination. Definite and clear signals following PCR amplification were obtained from all embryo samples. Accuracy of assays was determined by comparing results from a single cell with those of blastocyst stage embryos, thereby indicating that 1 or 2 blastomeres from a preimplantation buffalo embryo is sufficient for sex determination by PCR. No misidentification was observed within the embryo samples using nested (BuRY.I), primary (BRY.I) and multiplex (BuRYN.I; ZFX/ZFY) PCR, suggesting that this technique is a highly reliable method for sexing buffalo embryos.  相似文献   

10.
11.
Summary The divergence pattern of mammalian ZFY-related genes from human (ZFY and ZFX) and mouse (Zfy-1 and Zfx) was reexamined on the basis of nucleotide substitutions at the synonymous codon-alternating positions. It is possible to explain the unusual divergence pattern of the mammalian Y-linked ZF genes by interchromosomal gene conversion by X-linked ZF genes. Furthermore, the rates of evolution of mammalian X- and Y-linked ZF genes were shown to agree well with those expected from our model. Offprint requests to: T. Miyata  相似文献   

12.
We developed a single‐reaction test for identifying the sex of giant panda (Ailuropoda melanoleuca) targeted to co‐amplify homologous fragments with size polymorphism that located at zinc‐finger (ZF) intron 7 by using one pair of primers. This assay produced one sex‐specific fragment in females (XX genotypes) whereas two fragments were produced in males (XY genotypes). Indels (insertion/deletion) in intron 7 of Y‐linked allele provide a significant discrimination between ZFX and ZFY, thus the amplification products can be simply distinguished by agarose gel electrophoresis, exhibiting sex‐specific banding patterns (female, 354 bp; male, 354 bp, 135 bp). The new primer set was successfully tested on known‐sex giant pandas by using template DNA extracted from both blood and fecal samples. Cross‐species test was also performed, revealing that this assay could be applied to other Ursidae species. Zoo Biol 29:526–531, 2010. © 2009 Wiley‐Liss, Inc.  相似文献   

13.
Campomelic dysplasia (Cd) occurs combined with sex reversal resulting in XY females. The recent identification of candidate genes for sex determination/differentiation and of a sex determining region on the human Y chromosome prompted the authors to study these genes for mutations in patients with Cd and sex reversal. In a total of five cases, no evidence for a mutation in the genes SRY, ZFY, ZFX, MEA and some anonymous Y-linked sequences was found. In addition to Southern analysis, gene expression of ZFY, ZFX and MEA was found to be normal as well. It is concluded that sex reversal in this condition is due to mutation in a so far unidentified gene which may act secondary to the testis-determining factor (TDF).  相似文献   

14.
We have sequenced the partial exon of the zinc finger genes (ZFX and ZFY) in 5 hominoids, 2 Old World monkeys, 1 New World monkey, and 1 prosimian. Among these primate species, the percentage similarities of the nucleotide sequence of the ZFX gene were 96-100% and 91.2-99.7% for the ZFY gene. Of 397 sites in the ZFX and ZFY gene sequences, 20 for ZFX gene and 42 for ZFY gene were found to be variable. Substitution causes 1 amino acid change in ZFX, and 5 in ZFY, among 132 amino acids. The numbers of synonymous substitutions per site (Ks) between human and the chimpanzee, gorilla and orangutan for ZFY gene were 0.026, 0.033, and 0.085, respectively. In contrast, the Ks value between human and hominoid primates for the ZFX gene was 0.008 for each comparison. Comparison of the ZFX and ZFY genes revealed that the synonymous substitution levels were higher in hominoids than in other primates. The rates of synonymous substitution per site per year were higher in the ZFY exon than in the SRY exon, and higher in the ZFY exon than in the ZFY intron, in hominoid primates.  相似文献   

15.
An accurate, sensitive, and quick (approximately 3 h) method for determining the sex of ovine embryos was developed using polymerase chain reaction (PCR) primers derived from an ovine-specific Y-chromosome random amplified polymorphic DNA marker ( UcdO43 ). The accuracy and sensitivity of the assay were first tested using genomic DNA from 10 males and 10 females of five different sheep breeds, and then tested using serial dilutions of male-in-female DNA. The assay was 100% accurate in confirming the sex of the individuals and the ovine male-specific fragment was detected in dilutions containing as little as 10 pg of male DNA in 50 ng of female DNA. The assay was also confirmed to be specific for the ovine Y-chromosome as bovine, caprine, porcine, murine, and human DNA did not amplify. The ovine embryo sexing method is a duplex PCR system that also includes ZFY/ZFX primers. ZFY/ZFX provide an internal positive control for amplification as well as a means to confirm the results obtained with the UcdO43 primers. All embryo sexing results (36/36) from our method were in agreement with the ZFY/ZFX assay results. However, while our method requires an internal control to detect PCR failure, it has the advantages of not requiring nested PCR or restriction endonuclease digestion of the PCR product, and concerns about cross-species contamination are eliminated.  相似文献   

16.
Sex determination of bovine embryo blastomeres by fluorogenic probes   总被引:7,自引:0,他引:7  
One of the major challenges of using genetic information in marker assisted selection (MAS) is the detection of multiple marker loci from a small biopsy sample of a preimplantation stage embryo. The objective of this study was to develop a fast, nested, multiplex preamplification, polymerase chain reaction (PCR) method for the determination of sex in bovine embryo blastomeres. For this aim, ZFX/ZFY sequences were preamplified simultaneously with other genomic regions. The preamplification product was used as a template in an allelic discrimination assay, with nested primers and sex specific fluorogenic probes for ZFX and ZFY. Fluorogenic probes were used to eliminate the need for time consuming electrophoresis. Compared to sexing with Bovy/kappa-casein co-amplification method and other replicates from the same embryo, the accuracy of sexing with the use of fluorogenic probes after preamplification was 99% (112/113 blastomeres). The amplification efficiency was 96% (113/117 blastomeres).  相似文献   

17.
We developed a new PCR/RFLP system targeted to amplify and cut a segment of the ZFX/ZFY gene in non-invasive otter (Lutra lutra) samples. This assay produced one sex-specific fragment in females (XX genotypes) and two fragments in males (XY genotypes), and is intrinsically more reliable than alternative systems (e.g., SRY genes) that are based on the amplification of a single Y-linked fragment. The new primer pair correctly identified the sex of 23 DNA extracted from sexed otter tissues. This procedure was used to sex unknown DNA extracted from otter scats. A multi-tube approach led to identify the sex of 72/91 (79%) samples, using a minimum of two PCR replicates. This procedure is currently used in non-invasive genetic monitoring of Italian otter populations.  相似文献   

18.
A rapid and reliable method for sex determination of preimplantation-stage equine embryos has not been available. The aim of the present study was to find an enzyme which would distinguish sexes in the horse by finding a polymorphic restriction site between the ZFY and ZFX homologues amplified by the polymerase chain reaction (PCR). Altogether, 38 different restriction enzymes were tested using female and male DNA extracted from blood. The primers used for amplification were selected from conserved sequences between human ZFY and ZFX genes and mouse Zfy-1 and Zfy-2 genes. Nine enzymes cut the PCR product of approximately 450 basepairs, but only Bsm I yielded different banding patterns in female and male DNA. All blood samples were correctly diagnosed. To test the method on embryonic cells, 17 horse demi-embryos were obtained from expanding blastocysts 220 to 950 mum in diameter. Demi-embryos were further cut into 3 to 7 parallel samples which were analyzed individually to test the repeatability of the method. Eight of the original embryos were diagnosed as females and 9 as males. No misidentifications were observed within the embryonic samples, suggesting that this sexing method is highly reliable. This study provides a rapid and accurate method to sex horse embryos.  相似文献   

19.
20.
Summary Two 46,XY females with tandem duplications of an X short arm segment were studied by cytogenetic and Southern blot analysis. The results show that the duplicated segment in each case included the Xp21.2–Xp22.2 interval, resulting in a double dose of ZFX on the single active X chromosome. The results from our two cases, in conjunction with those reported by other workers, lead us to conclude that the duplication is the reason for the sex inversion. If ZFY and ZFX are indeed sex-determining gene loci, these findings favour a model of sex determination characterized by antagonistic interaction between these genes.  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