DNA甲基化和组蛋白修饰调节牛ZAR1基因表达的研究
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摘要
合子阻滞因子1(Zygote arrest 1,ZAR1)是卵母细胞特异的母性效应基,在胚胎早期发育的母型合子转变中起重要作用。牛ZAR1是单拷贝基因,位于第六号染色体,基因全长4126bp,编码序列为1487bp,具有四个外显子,3个内含子,包含1155bp的开放阅读框,编码387个氨基酸的前体蛋白。该蛋白具有在进化上保守的植物同源结构域PHD,表明ZAR1具有转录调节的作用,DNA甲基转移酶和组蛋白去乙酰化酶可能通过PHD等结构域抑制基因转录。ZAR1基因编码区有大量的回文序列,能够形成茎环结构和十字结构从而改变DNA构象,也可能直接和蛋白质相互作用。短回文序列具有核受体的DNA结合域,调节核受体的表达。该基因3′非翻译区含有UGUA序列和多聚脱腺苷化元件,与ZAR1蛋白的翻译有关。
ZAR1在小鼠卵巢中特异表达,在睾丸中不表达,ZAR1~(-/-)小鼠可以存活,但却是不育的。ZAR1~(-/-)小鼠在卵巢发育、卵母细胞发生和受精后的早期阶段虽然正常,但大多数来自ZAR1~(-/-)雌性的胚胎都阻滞于合子期,约有少于20%的胚胎到达2细胞期,但不能发育到4细胞。牛ZAR1在多种组织中表达,如心脏、睾丸、卵巢、骨骼肌,在胚胎发育不同阶段也有表达,有关该基因表达调控的表观遗传学机制还未见报道。本试验利用RT-PCR,亚硫酸盐测序PCR(bisulfite-sequencing PCR,BSP)、染色质免疫共沉淀(chromatinimmuno-precipitation assay,ChIP),等实验技术对成年牛组织中ZAR1基因表达情况及该基因5′调控区的DNA甲基化和组蛋白修饰进行研究,并利用实时定量PCR方法检测该基因在牛卵母细胞及体外受精胚胎中的mRNA表达的动态变化。
1、ZAR1基因在牛组织中的表达
利用RT-PCR检测成年牛心脏、肾脏、肝脏及睾丸中ZAR1基因mRNA的表达。结果显示ZAR1基因具有组织特异性表达:ZAR1基因在睾丸中表达较高,肾脏和心脏较低,肝脏中未检测到表达。
2、ZAR1基因DNA甲基化状态
DNA甲基化是调控基因表达的一种方式,主要发生在CpG位点的胞嘧啶的第5个碳原子上,DNA甲基化并不改变基因的碱基序列,而是通过调节基因的表达影响其功能。本试验选定ZAR1基因5′调控区中的11个CpG位点,以亚硫酸盐处理的基因组为模板,进行PCR并测序,检测CpG位点的甲基化状态,结果显示:心脏中的甲基化程度明显高于其他三种组织,说明该调控区DNA甲基化与牛ZAR1基因的组织特异性表达相关。
3、ZAR1基因组蛋白修饰状况
组蛋白修饰是另一种重要的表观遗传调节机制,组蛋白修饰通过改变染色体的构象从而影响转录因子与DNA的结合来调节基因表达。一般认为,H3乙酰化与基因活化有关,而H3K9甲基化与基因沉默有关。
本试验利用染色质免疫共沉淀方法对成年牛组织中ZAR1基因5′调控区的H3乙酰化、H3K9甲基化状况进行了检测。结果显示在检测的组织中心脏组织H3乙酰化程度较高,肝脏中较低。H3乙酰化较高可能与ZAR1在心脏中有表达相关;肝脏中乙酰化程度较低可能与肝脏中未检测到ZAR1的表达相关。本试验中没有在四种组织中检测到H3K9甲基化,H3K9甲基化与基因沉默相关,这一结果与ZAR1基因在所检测的牛的几种组织中的广泛表达相一致。有研究表明H3K9甲基化最主要作用靶标是卫星簇和编码区,而不是编码蛋白基因的启动子区,本研究所检测的调控区可能不是H3K9甲基化的主要修饰位点。
4、牛卵母细胞中ZAR1基因的表达
研究基因mRNA表达的方法有Northern杂交、实时荧光定量PCR及基因芯片等多种方法,其中实时荧光定量PCR技术不仅能进行初步定量研究,且与常规PCR相比特异性和灵敏性更强。本试验采用实时荧光定量PCR方法检测了ZAR1在牛卵母细胞成熟过程中五个时期(GV、PMI、MI、AI-TI、MII)的mRNA表达状况。ZAR1在生发泡和第一次减数分裂前中期表达量较高且有显著差异,而在第一次减数分裂中期、末期和第二次减数分裂中期表达量较低且没有显著差异。
5、牛体外受精胚胎中ZAR1基因的表达
ZAR1是胚胎发育的关键基因之一,检测ZAR1的表达状况为其功能研究提供一些依据。本试验应用实时荧光定量PCR方法检测了牛体外受精胚胎中ZAR1基因的表达状况。研究结果表明ZAR1在2-细胞、4-细胞、8-细胞中表达量较高,且有显著差异,而在桑椹胚和囊胚中表达量较低,且无显著差异。
综上所述,ZAR1是卵母细胞特异性母性效应基因,在胚胎早期发育的母型合子转变中起重要作用。本研究表明牛ZAR1基因的表达具有组织特异性,在心脏、肾脏、睾丸中有表达,肝脏中未检测到。对ZAR1基因5′调控区的DNA甲基化和组蛋白修饰的研究结果表明二者与ZAR1的组织特异性表达调控相关。ZAR1在牛卵母细胞成熟和早期胚胎发育中的表达具有动态变化,生发泡期表达量较高,随着减数分裂的进行表达量逐渐降低。值得注意的是,与MII期相比2-细胞期的ZAR1 mRNA水平明显升高,推测可能与母型合子转变有关。
Zygote arrest 1(ZAR1) gene plays a crucial role in maternal zygotic transition(MZT). Bovine ZAR1 is a single copy gene, located on chromosome 6, the full-length is 4126bp containing 1487bp, with 4 extrons and 3 introns, including 1155bp open reading frame, encoding precursor protein of 384 amino acid. The protein has an typical plant homeobox zing finger PHD motif involved in transcription regulation. It has been postulated that DNA methyltransferases interacting with HDACs repress transcription through PHD motif. ZAR1 coding region is proved including numerous palindrome repeats forming stem-loops and cruciform structure, which may interact directly with proteins. Short palindrome repeats are characteristic of DNA-binding domain of nuclear receptors. UGUA sequence and polyadenylation element were detected within 3'-UTR of ZAR1 mRNA, associated with ZAR1 translation regulation.
Mouse ZAR1 was expressed specifically in ovary but not in testis. Although ZAR1~(-/-) mice are viable and grossly normal, ZAR1~(-/-) females are infertile. Ovarian development and oogenesis and the early stages after fertilization seems unimpaired, but most embryos from ZAR1~(-/-) females arrest at the one-cell stage. Fewer than 20% of the embryos derived from ZAR1~(-/-) females develop to the two-cell stage and no embryos succeed to develop to the four-cell stage. ZAR1 was found in variant tissues including ovary, testis, skeletal muscle, and myocardium in cattle as well as in embryo development, which is not restricted in ovary. There is no report about the epigenetic regulation of the gene. In this study, RT-PCR and bisulfite-sequencing PCR and chromatin immuno-precipitation assay(ChIP) were used to detect the dynamic change of DNA methylation and histone modification in the regulatory region of bovine ZAR1 gene. Real time PCR was applied to detect ZAR1 dynamic expression pattern in bovine oocytes and IVF embryos.
1. expression of ZAR1 gene in bovine
RT-PCR was used to detect the expression of ZAR1 in bovine heart, kidney, liver and testis. The result showed that the expression of ZAR1 had a tissue-specific pattern that is abundant in testis, fewer in kidney and heart, but not detectable in liver.
2. DNA methylation status of ZAR1 gene
DNA methylation is one of the modes regulating gene expression, mainly methylated only at 5'cytosines in the CpG dinucleotide changed into guanosine. Without changing DNA sequence, DNA methylation influences the function of gene through regulating the gene expression. In this study 11 CpG sites in the 5' regulation region of ZAR1 gene were chosen to examine their DNA methylation status by bisulfite-sequencing PCR (BSP). The result showed that the methylation level in heart was significantly higher than in the other three tissues, demonstrating ZAR1 expression is associated with DNA methylation.
3. Histone modification pattern of ZAR1 gene
Histone modification is another important epigenetic regulation mechanism, which modulate gene expression through transforming the conformation of chromosome and thereby impact the interaction between the transcription factors and DNA. Generally H3 acetylation is related to gene activation, while H3K9 methylation is considered to be associated with gene silence.
ChIP was used to investigate the histone modification of 5' regulatory region in bovine heart, kidney, liver and testis. The result showed that acetylation level of H3 was higher in heart and lower in liver. The high level of H3 acetylation may lead to the expression in heart: the low level of H3 acetylation may be involved in the non-expression of ZAR1 in liver . H3K9 methylation was not detectable in tested tissues confirming H3K9me is associated with gene silencing and this result was consistent with the general expression of ZAR1 gene in bovine. It was reported that the major targets of H3K9 methylation were satellite domain and coding region, rather than the promoter region. Consequently, it is possible that H3K9 methylation was not the major modification form in the region detected in this study.
4. ZAR1 expression pattern in bovine oocytes
The methods of studying mRNA level include Northern blot, RT-PCR, real time PCR, gene chip and so on. Real time PCR is remarkable in qualitative analysis, exhibiting more specificity and sensitivity. Real time PCR was used to detect ZAR1 mRNA level during oocyte maturation(GV、PMI、MI、AI-TI、MII). The level of ZAR1 mRNA is high in GV and PMI phases with significant difference and lower in MI, AI-TI and MII phases without significant difference.
5. ZAR1 expression pattern in bovine IVF embryos
ZAR1 is a critical gene during embryo development and thereby it'll be helpful for the further research to detect ZAR1 expression pattern in bovine IVF embryos by real time PCR. The results showed that ZAR1 exhibited higher expression level in 2-cell, 4-cell and 8-cell with significant difference and lower level in morula and blastula without significant difference.
In conclusion, ZAR1 plays a critical role in maternal zygotic transition(MZT). This study showed the expression of ZAR1 had a tissue-specific pattern in bovine, which is detectable in heart, kidney and testis, but not in liver. The investigation of DNA methylation and histone modification status in the 5' regulation region of ZAR1 demonstrated both of them are correlated with the tissue-specific expression patten. The levels of ZAR1 mRNA present dynamic changes in oocyes maturation and early embryo development: ZAR1 mRNA is higher expressed in GV phase, decreases during oocyte maturation. It is notable that ZAR1 mRNA evidently increased in 2-cell stage, compared with MII, which is inferred that ZAR1 has a compact correlation with MZT.
ZAR1在小鼠卵巢中特异表达,在睾丸中不表达,ZAR1~(-/-)小鼠可以存活,但却是不育的。ZAR1~(-/-)小鼠在卵巢发育、卵母细胞发生和受精后的早期阶段虽然正常,但大多数来自ZAR1~(-/-)雌性的胚胎都阻滞于合子期,约有少于20%的胚胎到达2细胞期,但不能发育到4细胞。牛ZAR1在多种组织中表达,如心脏、睾丸、卵巢、骨骼肌,在胚胎发育不同阶段也有表达,有关该基因表达调控的表观遗传学机制还未见报道。本试验利用RT-PCR,亚硫酸盐测序PCR(bisulfite-sequencing PCR,BSP)、染色质免疫共沉淀(chromatinimmuno-precipitation assay,ChIP),等实验技术对成年牛组织中ZAR1基因表达情况及该基因5′调控区的DNA甲基化和组蛋白修饰进行研究,并利用实时定量PCR方法检测该基因在牛卵母细胞及体外受精胚胎中的mRNA表达的动态变化。
1、ZAR1基因在牛组织中的表达
利用RT-PCR检测成年牛心脏、肾脏、肝脏及睾丸中ZAR1基因mRNA的表达。结果显示ZAR1基因具有组织特异性表达:ZAR1基因在睾丸中表达较高,肾脏和心脏较低,肝脏中未检测到表达。
2、ZAR1基因DNA甲基化状态
DNA甲基化是调控基因表达的一种方式,主要发生在CpG位点的胞嘧啶的第5个碳原子上,DNA甲基化并不改变基因的碱基序列,而是通过调节基因的表达影响其功能。本试验选定ZAR1基因5′调控区中的11个CpG位点,以亚硫酸盐处理的基因组为模板,进行PCR并测序,检测CpG位点的甲基化状态,结果显示:心脏中的甲基化程度明显高于其他三种组织,说明该调控区DNA甲基化与牛ZAR1基因的组织特异性表达相关。
3、ZAR1基因组蛋白修饰状况
组蛋白修饰是另一种重要的表观遗传调节机制,组蛋白修饰通过改变染色体的构象从而影响转录因子与DNA的结合来调节基因表达。一般认为,H3乙酰化与基因活化有关,而H3K9甲基化与基因沉默有关。
本试验利用染色质免疫共沉淀方法对成年牛组织中ZAR1基因5′调控区的H3乙酰化、H3K9甲基化状况进行了检测。结果显示在检测的组织中心脏组织H3乙酰化程度较高,肝脏中较低。H3乙酰化较高可能与ZAR1在心脏中有表达相关;肝脏中乙酰化程度较低可能与肝脏中未检测到ZAR1的表达相关。本试验中没有在四种组织中检测到H3K9甲基化,H3K9甲基化与基因沉默相关,这一结果与ZAR1基因在所检测的牛的几种组织中的广泛表达相一致。有研究表明H3K9甲基化最主要作用靶标是卫星簇和编码区,而不是编码蛋白基因的启动子区,本研究所检测的调控区可能不是H3K9甲基化的主要修饰位点。
4、牛卵母细胞中ZAR1基因的表达
研究基因mRNA表达的方法有Northern杂交、实时荧光定量PCR及基因芯片等多种方法,其中实时荧光定量PCR技术不仅能进行初步定量研究,且与常规PCR相比特异性和灵敏性更强。本试验采用实时荧光定量PCR方法检测了ZAR1在牛卵母细胞成熟过程中五个时期(GV、PMI、MI、AI-TI、MII)的mRNA表达状况。ZAR1在生发泡和第一次减数分裂前中期表达量较高且有显著差异,而在第一次减数分裂中期、末期和第二次减数分裂中期表达量较低且没有显著差异。
5、牛体外受精胚胎中ZAR1基因的表达
ZAR1是胚胎发育的关键基因之一,检测ZAR1的表达状况为其功能研究提供一些依据。本试验应用实时荧光定量PCR方法检测了牛体外受精胚胎中ZAR1基因的表达状况。研究结果表明ZAR1在2-细胞、4-细胞、8-细胞中表达量较高,且有显著差异,而在桑椹胚和囊胚中表达量较低,且无显著差异。
综上所述,ZAR1是卵母细胞特异性母性效应基因,在胚胎早期发育的母型合子转变中起重要作用。本研究表明牛ZAR1基因的表达具有组织特异性,在心脏、肾脏、睾丸中有表达,肝脏中未检测到。对ZAR1基因5′调控区的DNA甲基化和组蛋白修饰的研究结果表明二者与ZAR1的组织特异性表达调控相关。ZAR1在牛卵母细胞成熟和早期胚胎发育中的表达具有动态变化,生发泡期表达量较高,随着减数分裂的进行表达量逐渐降低。值得注意的是,与MII期相比2-细胞期的ZAR1 mRNA水平明显升高,推测可能与母型合子转变有关。
Zygote arrest 1(ZAR1) gene plays a crucial role in maternal zygotic transition(MZT). Bovine ZAR1 is a single copy gene, located on chromosome 6, the full-length is 4126bp containing 1487bp, with 4 extrons and 3 introns, including 1155bp open reading frame, encoding precursor protein of 384 amino acid. The protein has an typical plant homeobox zing finger PHD motif involved in transcription regulation. It has been postulated that DNA methyltransferases interacting with HDACs repress transcription through PHD motif. ZAR1 coding region is proved including numerous palindrome repeats forming stem-loops and cruciform structure, which may interact directly with proteins. Short palindrome repeats are characteristic of DNA-binding domain of nuclear receptors. UGUA sequence and polyadenylation element were detected within 3'-UTR of ZAR1 mRNA, associated with ZAR1 translation regulation.
Mouse ZAR1 was expressed specifically in ovary but not in testis. Although ZAR1~(-/-) mice are viable and grossly normal, ZAR1~(-/-) females are infertile. Ovarian development and oogenesis and the early stages after fertilization seems unimpaired, but most embryos from ZAR1~(-/-) females arrest at the one-cell stage. Fewer than 20% of the embryos derived from ZAR1~(-/-) females develop to the two-cell stage and no embryos succeed to develop to the four-cell stage. ZAR1 was found in variant tissues including ovary, testis, skeletal muscle, and myocardium in cattle as well as in embryo development, which is not restricted in ovary. There is no report about the epigenetic regulation of the gene. In this study, RT-PCR and bisulfite-sequencing PCR and chromatin immuno-precipitation assay(ChIP) were used to detect the dynamic change of DNA methylation and histone modification in the regulatory region of bovine ZAR1 gene. Real time PCR was applied to detect ZAR1 dynamic expression pattern in bovine oocytes and IVF embryos.
1. expression of ZAR1 gene in bovine
RT-PCR was used to detect the expression of ZAR1 in bovine heart, kidney, liver and testis. The result showed that the expression of ZAR1 had a tissue-specific pattern that is abundant in testis, fewer in kidney and heart, but not detectable in liver.
2. DNA methylation status of ZAR1 gene
DNA methylation is one of the modes regulating gene expression, mainly methylated only at 5'cytosines in the CpG dinucleotide changed into guanosine. Without changing DNA sequence, DNA methylation influences the function of gene through regulating the gene expression. In this study 11 CpG sites in the 5' regulation region of ZAR1 gene were chosen to examine their DNA methylation status by bisulfite-sequencing PCR (BSP). The result showed that the methylation level in heart was significantly higher than in the other three tissues, demonstrating ZAR1 expression is associated with DNA methylation.
3. Histone modification pattern of ZAR1 gene
Histone modification is another important epigenetic regulation mechanism, which modulate gene expression through transforming the conformation of chromosome and thereby impact the interaction between the transcription factors and DNA. Generally H3 acetylation is related to gene activation, while H3K9 methylation is considered to be associated with gene silence.
ChIP was used to investigate the histone modification of 5' regulatory region in bovine heart, kidney, liver and testis. The result showed that acetylation level of H3 was higher in heart and lower in liver. The high level of H3 acetylation may lead to the expression in heart: the low level of H3 acetylation may be involved in the non-expression of ZAR1 in liver . H3K9 methylation was not detectable in tested tissues confirming H3K9me is associated with gene silencing and this result was consistent with the general expression of ZAR1 gene in bovine. It was reported that the major targets of H3K9 methylation were satellite domain and coding region, rather than the promoter region. Consequently, it is possible that H3K9 methylation was not the major modification form in the region detected in this study.
4. ZAR1 expression pattern in bovine oocytes
The methods of studying mRNA level include Northern blot, RT-PCR, real time PCR, gene chip and so on. Real time PCR is remarkable in qualitative analysis, exhibiting more specificity and sensitivity. Real time PCR was used to detect ZAR1 mRNA level during oocyte maturation(GV、PMI、MI、AI-TI、MII). The level of ZAR1 mRNA is high in GV and PMI phases with significant difference and lower in MI, AI-TI and MII phases without significant difference.
5. ZAR1 expression pattern in bovine IVF embryos
ZAR1 is a critical gene during embryo development and thereby it'll be helpful for the further research to detect ZAR1 expression pattern in bovine IVF embryos by real time PCR. The results showed that ZAR1 exhibited higher expression level in 2-cell, 4-cell and 8-cell with significant difference and lower level in morula and blastula without significant difference.
In conclusion, ZAR1 plays a critical role in maternal zygotic transition(MZT). This study showed the expression of ZAR1 had a tissue-specific pattern in bovine, which is detectable in heart, kidney and testis, but not in liver. The investigation of DNA methylation and histone modification status in the 5' regulation region of ZAR1 demonstrated both of them are correlated with the tissue-specific expression patten. The levels of ZAR1 mRNA present dynamic changes in oocyes maturation and early embryo development: ZAR1 mRNA is higher expressed in GV phase, decreases during oocyte maturation. It is notable that ZAR1 mRNA evidently increased in 2-cell stage, compared with MII, which is inferred that ZAR1 has a compact correlation with MZT.
引文
[1] Wu X, et al. Zygote arrest 1 (Zarl) is a novel maternal-effect gene critical for the oocyte-to-embryo transition. Nat Genet, 2003. 33(2): p. 187-191.
[2] Uzbekova, S., et al. Zygote arrest 1 gene in pig, cattle and human: evidence of different transcript variants in male and female germ cells. Reprod Biol Endocrinol, 2006. (4): p. 12-19.
[3] Goossens, K., et al. Selection of reference genes for quantitative real-time PCR in bovine preimplantation embryos. BMC Dev Biol, 2005. (5): p.27-29.
[4] Herman, J.G., et al. Methylation-specific PCR: a novel PCR assay for methylation status of CpG islands. Proc Natl Acad Sci U S A, 1996. 93(18): p.9821-9826.
[5] Roa, S.J., et al. Gene methylation patterns in digestive tumors. Rev Med Chil, 2008. 136(4): p.451-458.
[6] Young, L.E. and Beaujean N. DNA methylation in the preimplantation embryo: the differing stories of the mouse and sheep. Anim Reprod Sci, 2004.(8): p.61-78.
[7] Kaput, J. and Sneider T.W. Methylation of somatic vs germ cell DNAs analyzed by restriction endonuclease digestions. Nucleic Acids Res, 1979. 7(8): p.2303-2322.
[8] Farrell, W.E., et al. Methylation mechanisms in pituitary tumorigenesis. Endocr Relat Cancer, 1999. 6(4): p.437-447.
[9] Peterson, C.L. and Laniel M.A. Histones and histone modifications. Curr Biol, 2004. 14(14): p.546-551.
[10] Strahl, B.D. and Allis CD. The language of covalent histone modifications. Nature, 2000. 403(6765): p.41-45.
[1] Davey, C. A., Sargent, D. E, Luger, K. et al. Solvent mediated interactions in the structure of the nucleosome core particle at 1.9 a resolution. J Mol Biol, 2002, 319(5): p. 1097-1113.
[2] Kouzarides, T. Chromatin modifications and their function. Cell, 2007,128(4): p.693-705.
[3] Klose, R. J., Zhang, Y. Regulation of histone methylation by demethylimination and demethylation. Nat Rev Mol Cell Biol, 2007, 8(4): p.307-318.
[4] Surani, M. A., Hayashi, K., Hajkova, P. Genetic and epigenetic regulators of pluripotency. Cell, 2007,128(4): p.747-762.
[5] Agalioti, T., Chen, G., Thanos, D. Deciphering the transcriptional histone acetylation code for a human gene. Cell, 2002, 111(3): p.381-392.
[6] Grunstein, M. Histone acetylation in chromatin structure and transcription. Nature, 1997, 389(6649): p.349-352.
[7] Strahl, B. D., Allis, C. D. The language of covalent histone modifications. Nature, 2000, 403(6765): p.41-45.
[8] Kurdistani, S. K., Tavazoie, S., Grunstein, M. Mapping global histone acetylation patterns to gene expression. Cell, 2004, 117(6): p.721-733.
[9] Martin, C., Zhang, Y. The diverse functions of histone lysine methylation. Nat Rev Mol Cell Biol, 2005, 6(11):p.838-849.
[10] Jackson J P, Lindroth A M, Cao X, et al.Control of CpNpG DNA methylation by the KRYPTONITE histone H3 methyltransferase. Nature, 2002, (4): p.556-560.
[11] Bachman, K.E., et al. Histone modifications and silencing prior to DNA methylation of a tumor suppressor gene. Cancer Cell, 2003. 3(1): p.89-95.
[12] Sarraf, S.A. and Stancheva I. Methyl-CpG binding protein MBD1 couples histone H3 methylation at lysine 9 by SETDB1 to DNA replication and chromatin assembly. Mol Cell, 2004. 15(4):p.595-605.
[13] Ikura T, Ogryzko W, Grigoriev M, et al. Involvement of the TIP60 Histone Acetylase Complex in DNA Repair and Apoptosis. Cell, 2000, 102(4): p.463-473.
[14] Brownell, J. E., Zhou, J., Ranalli, T. et al. Tetrahymena histone acetyltransferaseA: a homolog to yeast Gcn5p linking histone acetylation to gene activation, Cell, 1996, 84(6): p.843-851.
[15] Cao, R., Wang, L., Wang, H. et al. Role of histone H3 lysine 27 methylation in Polycomb-group silencing. Science, 2002, 298(5595): p. 1039-1043.
[16] Rougeulle, C., Chaumeil, J., Sarma, K. et al. Differential histone H3 Lys-9 and Lys-27 methylation profiles on the X chromosome. Mol Cell Biol, 2004, 24(12): p.5475-5484.
[17] Arney, K.L., Fisher, A.G. Epigenetic aspects of differentiation, J Cell Sci, 2004, 117(19): p.4355-4363.
[18] Wu, J., et al. Diverse histone modifications on histone 3 lysine 9 and their relation to DNA methylation in specifying gene silencing. BMC Genomics, 2007. (8): p.131.
[1] Wu, X., et al. Zygote arrest 1 (Zarl) is an evolutionarily conserved gene expressed in vertebrate ovaries. Biol Reprod, 2003. 69(3): p.861-867.
[2] Telford, N.A., Watson A.J., and Schultz GA. Transition from maternal to embryonic control in early mammalian development: a comparison of several species. Mol Reprod Dev, 1990. 26(1): p.90-100.
[3] Evan, A.T., et al. The Role of Aerosols in the Evolution of Tropical North Atlantic Ocean Temperature Anomalies. Science, 2009. (3): p.225-231.
[4] Muller, K. and Wirth M. Real-time RT-PCR detection of retroviral contaminations of cells and cell lines. Cytotechnology, 2002. 38(1-3): p.147-153.
[5] Scipioni, A., et al. A SYBR Green RT-PCR assay in single tube to detect human and bovine noroviruses and control for inhibition. Virol J, 2008. (5): p.94-102.
[6] Thelie, A., et al. Differential regulation of abundance and deadenylation of maternal transcripts during bovine oocyte maturation in vitro and in vivo. BMC Dev Biol, 2007. (7): p.125.
[7] Goossens, K., et al. Selection of reference genes for quantitative real-time PCR in bovine preimplantation embryos. BMC Dev Biol, 2005.(6): p.883-893.
[8] Brevini, T.A., et al. Expression pattern of the maternal factor zygote arrest 1 (Zarl) in bovine tissues, oocytes, and embryos. Mol Reprod Dev, 2004. 69(4): p.375-380.
[9] Tong, Z.B., et al. Developmental expression and subcellular localization of mouse MATER, an oocyte-specific protein essential for early development. Endocrinology, 2004. 145(3): p. 1427-1434.
[10] Karen Goossens, Mario Van Poucke, Ann Van Soom, et al. Selection of reference genes for quantitative real-time PCR in bovine preimplantation embryos. BMC Developmental Biology 2005, (5): p.27-29.
[11] Wu X.M., Voveiros M.M., Eppig J.J., et al. Zygote arrest l(Zarl) is a novel maternal effect gene critical for the oocyte to embryo transition. Nat Ge net, 2003, (33): p.187 -191.
[12] Harper J.C., Pergament E., Delhanty J.D.A. Genetics of gametes and embryos. Eur J Obs Gyn Reprod Biol, 2004, (115): p.80-84.
[13] Latham K.E. Mechanisms and control of embryonic genome activation in mammalian embryos. Int Rev Cytol 1999, (193): p.71-124.
[1] Telford NA, Watson AJ, Schultz GA. Transition from maternal to embryonic control in early mammalian development: a comparison of several species. Mol Reprod Dev, 1990; 26(1); p.90-100.
[2] Zamir, E., Kam Z., and Yarden A.. Transcription-dependent induction of G1 phase during the zebra fish midblastula transition. Mol Cell Biol, 1997. 17(2): p.529-536.
[3] Grosshans, J., Muller H.A., and Wieschaus E. Control of cleavage cycles in Drosophila embryos by fruhstart. Dev Cell, 2003. 5(2): p.285-294.
[4] Ikegami, R., Hunter P., and Yager T.D. Developmental activation of the capability to undergo checkpoint-induced apoptosis in the early zebrafish embryo. Dev Biol, 1999. 209(2): p.409-433.
[5] Brauchle, M., Baumer K., and Gonczy P. Differential activation of the DNA replication checkpoint contributes to asynchrony of cell division in C. elegans embryos. Curr Biol, 2003. 13(10): p.819-827.
[6] Vigneault, C., et al. Transcription factor expression patterns in bovine in vitro-derived embryos prior to maternal-zygotic transition. Biol Reprod, 2004. 70(6): p.1701-1709.
[7] Tadros, W. and Lipshitz H.D. Setting the stage for development: mRNA translation and stability during oocyte maturation and egg activation in Drosophila. Dev Dyn, 2005. 232(3): p.593-608.
[8] Bashirullah, A., et al. Joint action of two RNA degradation pathways controls the timing of maternal transcript elimination at the midblastula transition in Drosophila melanogaster. Embo J, 1999. 18(9): p.2610-2620.
[9] Palancade, B., et al. Incomplete RNA polymerase II phosphorylation in Xenopus laevis early embryos. J Cell Sci, 2001. 114(Pt 13): p.2483-2489.
[10] Bellier, S., et al. Nuclear translocation and carboxyl-terminal domain phosphorylation of RNA polymerase II delineate the two phases of zygotic gene activation in mammalian embryos. Embo J, 1997. 16(20): p.6250-6262.
[11] Leclerc, V., Raisin S., and Leopold P. Dominant-negative mutants reveal a role for the Cdk7 kinase at the mid-blastula transition in Drosophila embryos. Embo J, 2000. 19(7): p. 1567-1575.
[12] Diaz, J., Pastor N., and Martinez-Mekler G. Role of a spatial distribution of IP3 receptors in the Ca~(2+) dynamics of the Xenopus embryo at the mid-blastula transition stage. Dev Dyn,
2005. 232(2):p.301-312.
[13] Kaneko, K.J. and DePamphilis M.L. Regulation of gene expression at the beginning of mammalian development and the TEAD family of transcription factors. Dev Genet, 1998. 22(1): p.43-55.
[14] Hair, A., et al. Control of gene expression in Xenopus early development. Dev Genet, 1998. 22(2): p.122-131.
[15] Flenniken, A.M. and Newrock K.M. H1 histone subtypes and subtype synthesis switches of normal and delobed embryos of Ilyanassa obsoleta. Dev Biol, 1987. 124(2): p.457-468.
[16] Wang, Z.F., et al. The mouse histone H1 genes: gene organization and differential regulation. JMol Biol, 1997. 271(1): p.124-138.
[17] Drabent, B., et al. Isolation of two murine H1 histone genes and chromosomal mapping of the H1 gene complement. Mamm Genome, 1995. 6(8): p.505-511.
[18] Kaye, P.L. and Church R.B. Uncoordinated synthesis of histones and DNA by mouse eggs and preimplantation embryos. J Exp Zool, 1983. 226(2): p.231-237.
[19] Brownell, J.E., et al. Tetrahymena histone acetyltransferase A: a homolog to yeast Gcn5p linking histone acetylation to gene activation. Cell, 1996. 84(6): p.843-851.
[20] Lee, D.Y., et al. A positive role for histone acetylation in transcription factor access to nucleosomal DNA. Cell, 1993. 72(1): p.73-84.
[21] Thompson, J.G., et al. Total protein content and protein synthesis within pre-elongation stage bovine embryos. Mol Reprod Dev, 1998. 50(2): p.139-145.
[22] Aoki, F., Worrad D.M., and Schultz R.M. Regulation of transcriptional activity during the first and second cell cycles in the preimplantation mouse embryo. Dev Biol, 1997. 181(2): p. 296-307.
[23] Clarke, H.J., McLay D.W., and Mohamed O.A. Linker histone transitions during mammalian oogenesis and embryogenesis. Dev Genet, 1998. 22(1): p.17-30.
[24] Tong, Z.B., et al. Mater, a maternal effect gene required for early embryonic development in mice. Nat Genet, 2000. 26(3): p.267-268.
[25] Pennetier, S., et al. Spatio-temporal expression of the germ cell marker genes MATER, ZARI, GDF9, BMP15,andVASA in adult bovine tissues, oocytes, and preimplantation embryos. Biol Reprod, 2004. 71(4): p. 1359-1366.
[26] Itoh, S., et al.,Enhanced bone regeneration by electrical polarization of hydroxyapatite. Artif Organs, 2006. 30(11): p.863-869.
[27] Moorde, C.H. and Richter J.D. Cytoplasmic polyadenylation elements mediate masking and unmasking of cyclin B1 mRNA. Embo J, 1999. 18(8): p.2294-2303.
[28] Stutz, A., et al. Masking, unmasking, and regulated polyadenylation cooperate in the translational control of a dormant mRNA in mouse oocytes. Genes Dev, 1998. 12(16): p. 2535-2248.
[29] Tong, Z.B., et al. Developmental expression and subcellular localization of mouse MATER, an oocyte-specific protein essential for early development. Endocrinology, 2004. 145(3): p. 1427-1434.
[30] Barnett, D.K., Kimura J., and Bavister B.D. Translocation of active mitochondria during hamster preimplantation embryo development studied by confocal laser scanning microscopy. Dev Dyn, 1996. 205(1): p.64-72.
[31] Dundr, M. and Misteli T. Functional architecture in the cell nucleus. Biochem J, 2001. 356(Pt 2):p.297-310.
[32] Flach, G., et al. The transition from maternal to embryonic control in the 2-cell mouse embryo. Embo J, 1982. 1(6): p.681-686.
[33] Schultz, R.M. Regulation of zygotic gene activation in the mouse. Bioessays, 1993. 15(8): p. 531-538.
[34] Saraste, M., Sibbald P.R., and Wittinghofer A. The P-loop~a common motif in ATP- and GTP-binding proteins. Trends Biochem Sci, 1990. 15(11): p.430-434.
[35] Tong, Z.B., Nelson L.M., and Dean J. Mater encodes a maternal protein in mice with a leucine-rich repeat domain homologous to porcine ribonuclease inhibitor. Mamm Genome, 2000. 11(4): p.281-287.
[36] Wu X, Viveiros MM, Eppig JJ, Bai Y, Fitzpatrick SL, and Matzuk MM. Zygote arrest 1(Zarl) is a novel maternal effect gene critical for the oocyte-to-embryo transition. NatGenet 2003; 33(2): p.187-191.
[37] Uzbekova, S., et al. Zygote arrest 1 gene in pig, cattle and human: evidence of different transcript variants in male and female germ cells. Reprod Biol Endocrinol, 2006. (4): p. 12-18.
[38] Wu, X., et al. Zygote arrest 1 (Zarl) is an evolutionarily conserved gene expressed in vertebrate ovaries. Biol Reprod, 2003. 69(3): p.861-867.
[2] Uzbekova, S., et al. Zygote arrest 1 gene in pig, cattle and human: evidence of different transcript variants in male and female germ cells. Reprod Biol Endocrinol, 2006. (4): p. 12-19.
[3] Goossens, K., et al. Selection of reference genes for quantitative real-time PCR in bovine preimplantation embryos. BMC Dev Biol, 2005. (5): p.27-29.
[4] Herman, J.G., et al. Methylation-specific PCR: a novel PCR assay for methylation status of CpG islands. Proc Natl Acad Sci U S A, 1996. 93(18): p.9821-9826.
[5] Roa, S.J., et al. Gene methylation patterns in digestive tumors. Rev Med Chil, 2008. 136(4): p.451-458.
[6] Young, L.E. and Beaujean N. DNA methylation in the preimplantation embryo: the differing stories of the mouse and sheep. Anim Reprod Sci, 2004.(8): p.61-78.
[7] Kaput, J. and Sneider T.W. Methylation of somatic vs germ cell DNAs analyzed by restriction endonuclease digestions. Nucleic Acids Res, 1979. 7(8): p.2303-2322.
[8] Farrell, W.E., et al. Methylation mechanisms in pituitary tumorigenesis. Endocr Relat Cancer, 1999. 6(4): p.437-447.
[9] Peterson, C.L. and Laniel M.A. Histones and histone modifications. Curr Biol, 2004. 14(14): p.546-551.
[10] Strahl, B.D. and Allis CD. The language of covalent histone modifications. Nature, 2000. 403(6765): p.41-45.
[1] Davey, C. A., Sargent, D. E, Luger, K. et al. Solvent mediated interactions in the structure of the nucleosome core particle at 1.9 a resolution. J Mol Biol, 2002, 319(5): p. 1097-1113.
[2] Kouzarides, T. Chromatin modifications and their function. Cell, 2007,128(4): p.693-705.
[3] Klose, R. J., Zhang, Y. Regulation of histone methylation by demethylimination and demethylation. Nat Rev Mol Cell Biol, 2007, 8(4): p.307-318.
[4] Surani, M. A., Hayashi, K., Hajkova, P. Genetic and epigenetic regulators of pluripotency. Cell, 2007,128(4): p.747-762.
[5] Agalioti, T., Chen, G., Thanos, D. Deciphering the transcriptional histone acetylation code for a human gene. Cell, 2002, 111(3): p.381-392.
[6] Grunstein, M. Histone acetylation in chromatin structure and transcription. Nature, 1997, 389(6649): p.349-352.
[7] Strahl, B. D., Allis, C. D. The language of covalent histone modifications. Nature, 2000, 403(6765): p.41-45.
[8] Kurdistani, S. K., Tavazoie, S., Grunstein, M. Mapping global histone acetylation patterns to gene expression. Cell, 2004, 117(6): p.721-733.
[9] Martin, C., Zhang, Y. The diverse functions of histone lysine methylation. Nat Rev Mol Cell Biol, 2005, 6(11):p.838-849.
[10] Jackson J P, Lindroth A M, Cao X, et al.Control of CpNpG DNA methylation by the KRYPTONITE histone H3 methyltransferase. Nature, 2002, (4): p.556-560.
[11] Bachman, K.E., et al. Histone modifications and silencing prior to DNA methylation of a tumor suppressor gene. Cancer Cell, 2003. 3(1): p.89-95.
[12] Sarraf, S.A. and Stancheva I. Methyl-CpG binding protein MBD1 couples histone H3 methylation at lysine 9 by SETDB1 to DNA replication and chromatin assembly. Mol Cell, 2004. 15(4):p.595-605.
[13] Ikura T, Ogryzko W, Grigoriev M, et al. Involvement of the TIP60 Histone Acetylase Complex in DNA Repair and Apoptosis. Cell, 2000, 102(4): p.463-473.
[14] Brownell, J. E., Zhou, J., Ranalli, T. et al. Tetrahymena histone acetyltransferaseA: a homolog to yeast Gcn5p linking histone acetylation to gene activation, Cell, 1996, 84(6): p.843-851.
[15] Cao, R., Wang, L., Wang, H. et al. Role of histone H3 lysine 27 methylation in Polycomb-group silencing. Science, 2002, 298(5595): p. 1039-1043.
[16] Rougeulle, C., Chaumeil, J., Sarma, K. et al. Differential histone H3 Lys-9 and Lys-27 methylation profiles on the X chromosome. Mol Cell Biol, 2004, 24(12): p.5475-5484.
[17] Arney, K.L., Fisher, A.G. Epigenetic aspects of differentiation, J Cell Sci, 2004, 117(19): p.4355-4363.
[18] Wu, J., et al. Diverse histone modifications on histone 3 lysine 9 and their relation to DNA methylation in specifying gene silencing. BMC Genomics, 2007. (8): p.131.
[1] Wu, X., et al. Zygote arrest 1 (Zarl) is an evolutionarily conserved gene expressed in vertebrate ovaries. Biol Reprod, 2003. 69(3): p.861-867.
[2] Telford, N.A., Watson A.J., and Schultz GA. Transition from maternal to embryonic control in early mammalian development: a comparison of several species. Mol Reprod Dev, 1990. 26(1): p.90-100.
[3] Evan, A.T., et al. The Role of Aerosols in the Evolution of Tropical North Atlantic Ocean Temperature Anomalies. Science, 2009. (3): p.225-231.
[4] Muller, K. and Wirth M. Real-time RT-PCR detection of retroviral contaminations of cells and cell lines. Cytotechnology, 2002. 38(1-3): p.147-153.
[5] Scipioni, A., et al. A SYBR Green RT-PCR assay in single tube to detect human and bovine noroviruses and control for inhibition. Virol J, 2008. (5): p.94-102.
[6] Thelie, A., et al. Differential regulation of abundance and deadenylation of maternal transcripts during bovine oocyte maturation in vitro and in vivo. BMC Dev Biol, 2007. (7): p.125.
[7] Goossens, K., et al. Selection of reference genes for quantitative real-time PCR in bovine preimplantation embryos. BMC Dev Biol, 2005.(6): p.883-893.
[8] Brevini, T.A., et al. Expression pattern of the maternal factor zygote arrest 1 (Zarl) in bovine tissues, oocytes, and embryos. Mol Reprod Dev, 2004. 69(4): p.375-380.
[9] Tong, Z.B., et al. Developmental expression and subcellular localization of mouse MATER, an oocyte-specific protein essential for early development. Endocrinology, 2004. 145(3): p. 1427-1434.
[10] Karen Goossens, Mario Van Poucke, Ann Van Soom, et al. Selection of reference genes for quantitative real-time PCR in bovine preimplantation embryos. BMC Developmental Biology 2005, (5): p.27-29.
[11] Wu X.M., Voveiros M.M., Eppig J.J., et al. Zygote arrest l(Zarl) is a novel maternal effect gene critical for the oocyte to embryo transition. Nat Ge net, 2003, (33): p.187 -191.
[12] Harper J.C., Pergament E., Delhanty J.D.A. Genetics of gametes and embryos. Eur J Obs Gyn Reprod Biol, 2004, (115): p.80-84.
[13] Latham K.E. Mechanisms and control of embryonic genome activation in mammalian embryos. Int Rev Cytol 1999, (193): p.71-124.
[1] Telford NA, Watson AJ, Schultz GA. Transition from maternal to embryonic control in early mammalian development: a comparison of several species. Mol Reprod Dev, 1990; 26(1); p.90-100.
[2] Zamir, E., Kam Z., and Yarden A.. Transcription-dependent induction of G1 phase during the zebra fish midblastula transition. Mol Cell Biol, 1997. 17(2): p.529-536.
[3] Grosshans, J., Muller H.A., and Wieschaus E. Control of cleavage cycles in Drosophila embryos by fruhstart. Dev Cell, 2003. 5(2): p.285-294.
[4] Ikegami, R., Hunter P., and Yager T.D. Developmental activation of the capability to undergo checkpoint-induced apoptosis in the early zebrafish embryo. Dev Biol, 1999. 209(2): p.409-433.
[5] Brauchle, M., Baumer K., and Gonczy P. Differential activation of the DNA replication checkpoint contributes to asynchrony of cell division in C. elegans embryos. Curr Biol, 2003. 13(10): p.819-827.
[6] Vigneault, C., et al. Transcription factor expression patterns in bovine in vitro-derived embryos prior to maternal-zygotic transition. Biol Reprod, 2004. 70(6): p.1701-1709.
[7] Tadros, W. and Lipshitz H.D. Setting the stage for development: mRNA translation and stability during oocyte maturation and egg activation in Drosophila. Dev Dyn, 2005. 232(3): p.593-608.
[8] Bashirullah, A., et al. Joint action of two RNA degradation pathways controls the timing of maternal transcript elimination at the midblastula transition in Drosophila melanogaster. Embo J, 1999. 18(9): p.2610-2620.
[9] Palancade, B., et al. Incomplete RNA polymerase II phosphorylation in Xenopus laevis early embryos. J Cell Sci, 2001. 114(Pt 13): p.2483-2489.
[10] Bellier, S., et al. Nuclear translocation and carboxyl-terminal domain phosphorylation of RNA polymerase II delineate the two phases of zygotic gene activation in mammalian embryos. Embo J, 1997. 16(20): p.6250-6262.
[11] Leclerc, V., Raisin S., and Leopold P. Dominant-negative mutants reveal a role for the Cdk7 kinase at the mid-blastula transition in Drosophila embryos. Embo J, 2000. 19(7): p. 1567-1575.
[12] Diaz, J., Pastor N., and Martinez-Mekler G. Role of a spatial distribution of IP3 receptors in the Ca~(2+) dynamics of the Xenopus embryo at the mid-blastula transition stage. Dev Dyn,
2005. 232(2):p.301-312.
[13] Kaneko, K.J. and DePamphilis M.L. Regulation of gene expression at the beginning of mammalian development and the TEAD family of transcription factors. Dev Genet, 1998. 22(1): p.43-55.
[14] Hair, A., et al. Control of gene expression in Xenopus early development. Dev Genet, 1998. 22(2): p.122-131.
[15] Flenniken, A.M. and Newrock K.M. H1 histone subtypes and subtype synthesis switches of normal and delobed embryos of Ilyanassa obsoleta. Dev Biol, 1987. 124(2): p.457-468.
[16] Wang, Z.F., et al. The mouse histone H1 genes: gene organization and differential regulation. JMol Biol, 1997. 271(1): p.124-138.
[17] Drabent, B., et al. Isolation of two murine H1 histone genes and chromosomal mapping of the H1 gene complement. Mamm Genome, 1995. 6(8): p.505-511.
[18] Kaye, P.L. and Church R.B. Uncoordinated synthesis of histones and DNA by mouse eggs and preimplantation embryos. J Exp Zool, 1983. 226(2): p.231-237.
[19] Brownell, J.E., et al. Tetrahymena histone acetyltransferase A: a homolog to yeast Gcn5p linking histone acetylation to gene activation. Cell, 1996. 84(6): p.843-851.
[20] Lee, D.Y., et al. A positive role for histone acetylation in transcription factor access to nucleosomal DNA. Cell, 1993. 72(1): p.73-84.
[21] Thompson, J.G., et al. Total protein content and protein synthesis within pre-elongation stage bovine embryos. Mol Reprod Dev, 1998. 50(2): p.139-145.
[22] Aoki, F., Worrad D.M., and Schultz R.M. Regulation of transcriptional activity during the first and second cell cycles in the preimplantation mouse embryo. Dev Biol, 1997. 181(2): p. 296-307.
[23] Clarke, H.J., McLay D.W., and Mohamed O.A. Linker histone transitions during mammalian oogenesis and embryogenesis. Dev Genet, 1998. 22(1): p.17-30.
[24] Tong, Z.B., et al. Mater, a maternal effect gene required for early embryonic development in mice. Nat Genet, 2000. 26(3): p.267-268.
[25] Pennetier, S., et al. Spatio-temporal expression of the germ cell marker genes MATER, ZARI, GDF9, BMP15,andVASA in adult bovine tissues, oocytes, and preimplantation embryos. Biol Reprod, 2004. 71(4): p. 1359-1366.
[26] Itoh, S., et al.,Enhanced bone regeneration by electrical polarization of hydroxyapatite. Artif Organs, 2006. 30(11): p.863-869.
[27] Moorde, C.H. and Richter J.D. Cytoplasmic polyadenylation elements mediate masking and unmasking of cyclin B1 mRNA. Embo J, 1999. 18(8): p.2294-2303.
[28] Stutz, A., et al. Masking, unmasking, and regulated polyadenylation cooperate in the translational control of a dormant mRNA in mouse oocytes. Genes Dev, 1998. 12(16): p. 2535-2248.
[29] Tong, Z.B., et al. Developmental expression and subcellular localization of mouse MATER, an oocyte-specific protein essential for early development. Endocrinology, 2004. 145(3): p. 1427-1434.
[30] Barnett, D.K., Kimura J., and Bavister B.D. Translocation of active mitochondria during hamster preimplantation embryo development studied by confocal laser scanning microscopy. Dev Dyn, 1996. 205(1): p.64-72.
[31] Dundr, M. and Misteli T. Functional architecture in the cell nucleus. Biochem J, 2001. 356(Pt 2):p.297-310.
[32] Flach, G., et al. The transition from maternal to embryonic control in the 2-cell mouse embryo. Embo J, 1982. 1(6): p.681-686.
[33] Schultz, R.M. Regulation of zygotic gene activation in the mouse. Bioessays, 1993. 15(8): p. 531-538.
[34] Saraste, M., Sibbald P.R., and Wittinghofer A. The P-loop~a common motif in ATP- and GTP-binding proteins. Trends Biochem Sci, 1990. 15(11): p.430-434.
[35] Tong, Z.B., Nelson L.M., and Dean J. Mater encodes a maternal protein in mice with a leucine-rich repeat domain homologous to porcine ribonuclease inhibitor. Mamm Genome, 2000. 11(4): p.281-287.
[36] Wu X, Viveiros MM, Eppig JJ, Bai Y, Fitzpatrick SL, and Matzuk MM. Zygote arrest 1(Zarl) is a novel maternal effect gene critical for the oocyte-to-embryo transition. NatGenet 2003; 33(2): p.187-191.
[37] Uzbekova, S., et al. Zygote arrest 1 gene in pig, cattle and human: evidence of different transcript variants in male and female germ cells. Reprod Biol Endocrinol, 2006. (4): p. 12-18.
[38] Wu, X., et al. Zygote arrest 1 (Zarl) is an evolutionarily conserved gene expressed in vertebrate ovaries. Biol Reprod, 2003. 69(3): p.861-867.