Xotch, the Xenopus homolog of Drosophila notch

During the development of a vertebrate embryo, cell fate is determined by inductive signals passing between neighboring tissues. Such determinative interactions have been difficult to characterize fully without knowledge of the molecular mechanisms involved. Mutations of Drosophila and the nematode Caenorhabditis elegans have been isolated that define a family of related gene products involved in similar types of cellular inductions. One of these genes, the Notch gene from Drosophila, is involved with cell fate choices in the neurogenic region of the blastoderm, in the developing nervous system, and in the eye-antennal imaginal disc. Complementary DNA clones were isolated from Xenopus embryos with Notch DNA in order to investigate whether cell-cell interactions in vertebrate embryos also depend on Notch-like molecules. This approach identified a Xenopus molecule, Xotch, which is remarkably similar to Drosophila Notch in both structure and developmental expression.     

Retinoic acid controls the bilateral symmetry of somite formation in the mouse embryo

A striking characteristic of vertebrate embryos is their bilaterally symmetric body plan, which is particularly obvious at the level of the somites and their derivatives such as the vertebral column. Segmentation of the presomitic mesoderm must therefore be tightly coordinated along the left and right embryonic sides. We show that mutant mice defective for retinoic acid synthesis exhibit delayed somite formation on the right side. Asymmetric somite formation correlates with a left-right desynchronization of the segmentation clock oscillations. These data implicate retinoic acid as an endogenous signal that maintains the bilateral synchrony of mesoderm segmentation, and therefore controls bilateral symmetry, in vertebrate embryos.     

Induction of mesoderm by a viral oncogene in early Xenopus embryos

During frog embryogenesis, mesoderm is specified in the equatorial region of the early embryo by a signal from the vegetal hemisphere. Prospective ectodermal cells dissected from the animal hemisphere can be respecified to form mesodermal tissues by recombination with vegetal tissue or by treatment with any of several polypeptide growth factors or growth factor-like molecules. Together with the discovery that several developmental mutations in Drosophila are in genes with significant homology to mammalian mitogens and oncogenes, these observations suggest that early developmental signals may use similar transduction pathways to mitogenic signals characterized in cultured mammalian cells. Whether mesoderm can be induced by activation of intracellular signal transduction pathways implicated in mitogenesis and oncogenesis has been investigated with the viral oncogene polyoma middle T. Microinjection of middle T messenger RNA into early embryos results in the respecification of isolated prospective ectodermal tissue to form characteristic mesodermal structures. Middle T in frog blastomeres appears to associate with cellular activities similar to those observed in polyoma-transformed mouse cells, and transformation-defective middle T mutants fail to induce mesoderm. These results suggest that early inductive signals and mitogenic and oncogenic stimuli may share common signal transduction pathways.     

不同培养液对牛体细胞核移植胚胎发育的影响

旨在为牛体细胞克隆胚胎体外培养体系的建立奠定基础。利用体细胞核移植技术生产牛重构胚,将重构胚随机分成2组,分别用mSOF和G1.5/G2.5培养,统计2种培养液对牛核移植克隆胚胎发育的影响,用TUNEL荧光检测系统检测2个培养组胚胎细胞凋亡情况,同时用Real time RT-PCR检测热应激蛋白基因Hsp70和凋亡相关基因Bax在2个培养组的相对表达水平。两组的卵裂率分别为85.6%和87.0%、囊胚率分别为22.9%和27.1%,差异都不显著(P>0.05),但G1.5/G2.5组的8-细胞形成率显著高于mSOF组(P<0.05);mSOF组凋亡率为12.1%±1.9%,显著高于G1.5/G2.5组(P<0.05),G1.5/G2.5组的Hsp70和Bax相对表达量显著低于mSOF组(P<0.05)。结果表明,G1.5/G2.5培养液更有利于牛体细胞核移植胚胎的体外发育。     

Cell recognition during neuronal development

Insect embryos, with their relatively simple nervous systems, provide a model system with which to study the cellular and molecular mechanisms underlying cell recognition during neuronal development. Such an approach can take advantage of the accessible cells of the grasshopper embryo and the accessible genes of Drosophila. The growth cones of identified neurons express selective affinities for specific axonal surfaces; such specificities give rise to the stereotyped patterns of selective fasciculation common to both species. These and other results suggest that early in development cell lineage and cell interactions lead to the differential expression of cell recognition molecules on the surfaces of small subsets of embryonic neurons whose axons selectively fasciculate with one another. Monoclonal antibodies reveal surface molecules in the Drosophila embryo whose expression correlates with this prediction. It should now be possible to isolate the genes encoding these potential cell recognition molecules and to test their function through the use of molecular genetic approaches in Drosophila.     

植物体细胞胚胎发生及其分子调控机制研究进展

植物的每个细胞都包含着该物种的全部遗传信息,具备发育成完整植株的遗传能力,这被称为植物细胞的全能性。体细胞胚胎(体胚)发生是指在没有受精的情况下,由体细胞或营养细胞发育成胚胎,是诱导植物细胞全能性的一种形式。体胚发生在种质资源保存、种苗生产、分子育种和植物基础研究等方面都有着广泛的应用,已成为重要的植物生物技术工具和研究平台。多年来的分子遗传学研究表明:体胚发生受到由众多转录因子、激素信号途径及表观遗传修饰等构成的复杂网络的调控。本研究概述了植物体胚发生的途径,并重点综述了体胚发生关键基因的功能与调控机制、体胚发生的表观遗传修饰以及体胚发生关键基因在基因工程中的应用。随着研究的深入和新技术的出现,体胚发生过程中涉及的代谢组分动态变化、转录调控、激素信号转导与表观遗传调控等复杂生物学过程有望得到更深入地阐释,将更进一步地解析植物体胚发生的分子调控机制。此外,利用体胚发生关键基因的功能与调控机制,开发更高效的体胚诱导和遗传转化方法,有望为更多植物的基因功能研究和遗传改良提供新的思路和技术。参81     

Functional genomic analysis of the Wnt-wingless signaling pathway

The Wnt-Wingless (Wg) pathway is one of a core set of evolutionarily conserved signaling pathways that regulates many aspects of metazoan development. Aberrant Wnt signaling has been linked to human disease. In the present study, we used a genomewide RNA interference (RNAi) screen in Drosophila cells to screen for regulators of the Wnt pathway. We identified 238 potential regulators, which include known pathway components, genes with functions not previously linked to this pathway, and genes with no previously assigned functions. Reciprocal-Best-Blast analyses reveal that 50% of the genes identified in the screen have human orthologs, of which approximately 18% are associated with human disease. Functional assays of selected genes from the cell-based screen in Drosophila, mammalian cells, and zebrafish embryos demonstrated that these genes have evolutionarily conserved functions in Wnt signaling. High-throughput RNAi screens in cultured cells, followed by functional analyses in model organisms, prove to be a rapid means of identifying regulators of signaling pathways implicated in development and disease.     

枣幼胚体细胞胚发生发育的组织学观察

为深入研究枣体细胞胚胎发生的形态建成过程,以小梨枣幼胚为材料,采用苏木精-伊红法染色和石蜡切片技术对体细胞胚发生、发育进行了组织学观察。结果表明：胚型愈伤组织细胞体积小,细胞核大,细胞质浓,细胞排列紧密；而非胚性愈伤组织细胞体积大,细胞核小,细胞质稀薄。体细胞胚产生于幼胚愈伤组织的表层细胞或内部细胞,且发生不同步,其发育过程与合子胚发育过程相似,即经历球形期、心形期和鱼雷期,最终形成子叶形胚。     

番木瓜美中红品种体胚转化体系的建立

利用已构建的番木瓜环斑病毒(Papaya ringspot virus,PRSV)基因,通过农杆菌介导法,建立美中红番木瓜(Carica papaya L.)体胚的转化体系。结果表明:共培养时,当农杆菌EHA105携带pCAMBIA2300质粒时,农杆菌的浓度(OD600 nm)应小于或等于0.1,其共培养后抑制农杆菌的羧苄青霉素浓度应以750 mg/L为宜;当农杆菌EHA105携带pBI121质粒时,则农杆菌的浓度(OD600 nm)应小于或等于0.8,其共培养后抑制农杆菌的羧苄青霉素的浓度则为500 mg/L;加上滤纸处理,抗生素洗涤以及洗涤后的干燥处理,可完全抑制体胚表面的农杆菌生长。对再生的番木瓜体胚进行PCR检测,结果表明PRSV基因已转入体胚中,但还需要成苗后的进一步检测。     

Shaping the vertebrate body plan by polarized embryonic cell movements

Polarized cell movements shape the major features of the vertebrate body plan during development. The head-to-tail body axis of vertebrates is elongated in embryonic stages by "convergent extension" tissue movements. During these movements cells intercalate between one another transverse to the elongating body axis to form a narrower, longer array. Recent discoveries show that these polarized cell movements are controlled by homologs of genes that control the polarity of epithelial cells in the developing wing and eye of the fruit fly, Drosophila.     

蜜蜂发育相关基因研究

蜜蜂是一种重要的授粉昆虫,也是研究人类疾病及社会行为的模式生物之一。由其基因组测序可知,蜜蜂基因组中(A+T)及CpG含量较高,其昼夜节律、RNAi和DNA甲基化基因更类似脊椎动物。先天免疫、表皮蛋白和味觉受体基因较少,气味受体基因较多。蜜蜂早期发育途径中的一些基因与果蝇的相似,功能却显著不同。本文综述了蜜蜂卵、幼虫、蛹和成虫期发育相关基因及其研究进展。     

棉花体细胞胚萌发及植株再生的研究

对棉花体细胞胚萌发规律及其影响因素进行研究结果,棉花体细胞胚的葫发可分为3种类型:①先出根,后出芽;②先出茅,后出根;③根、芽平衡生长,且3种类型所占的比例不同.不同基础培养基、不同激素种类和浓度,以及不同蔗糖浓度对棉花体细胞胚的萌发和植株再生的影响不同.BT培养基有利于根的发育,MS培养基有利于芽的发育,而SH培养基则有利于根、芽的平衡生长.KT的使用显著降低了棉花体细胞胚的生根率、生芽率、葫发率及再生植株率.低浓度的NAA有利于胚的葫发和再生植株,其最佳浓度为0.1 mg/L;MS培养基中添加0.1mg/L NAA,可使芽的分化率提高83.3%,再生植株率提高50.0%;而高浓度的NAA又抑制了芽的发育.在棉花体细胞胚萌发中蔗糖的最佳使用量为20g/L.     

南极独角雪冰鱼(Chionodraco hamatus)miR-204对斑马鱼胚胎血管发育的作用机制研究

南极独角雪冰鱼（Chionodraco hamatus）是目前已知的唯一缺乏血红蛋白、且不具有功能性血红细胞的脊椎动物。与相同生境下的其他南极鱼相比，其血管明显变粗，毛细血管增多。前期对南极独角雪冰鱼与近缘种伯氏肩孔南极鱼（Trematomus bernacchii）的血管microRNAs表达进行对比，发现南极独角雪冰鱼的miR-204的表达与伯氏肩孔南极鱼间存在显著差异，推测miR-204极有可能参与了南极冰鱼血管发育的调控。为证明此假设，运用斑马鱼（Barchydanio rerio）这一模式生物，利用斑马鱼胚胎显微注射技术、靶基因预测、双荧光素酶报告系统等一系列分子生物学手段对miR-204在斑马鱼血管发育中的调控作用进行探究。结果显示，抑制miR-204表达能使斑马鱼胚胎躯干部毛细血管增生。荧光素酶活性分析表明，miR-204可以显著降低trib3的表达，同时在斑马鱼胚胎中共注射miR-204和trib3后的GFP萤光强度显著降低，GFP蛋白量减少，说明miR-204对trib3有抑制作用。trib3最初发现是抑制果蝇胚胎和生殖细胞有丝分裂的假激酶，后续研究发现其参与动脉粥样硬化的调控以及糖尿病血管纤维化。综上，推测miR-204可能通过抑制trib3的表达进而调控毛细血管的增生。     

抗性湿地松体胚的发育、成熟与萌发

为加快抗松针褐斑病湿地松体细胞胚胎发生技术的产业化应用进程,本研究以肌醇浓度、脱落酸(ABA)的添加方式、基本培养基、ABA与聚乙二醇(PEG6000)互作、PEG6000及琼脂糖种类及浓度、活性炭(AC)、ABA、液体悬浮培养等因素对抗性湿地松体胚的发育、成熟与萌发的影响进行研究。结果表明:添加8g/L的肌醇为宜,过高浓度的肌醇则不利于湿地松体胚的发育；ABA可以采用高压灭菌；基本培养基、ABA、PEG6000互作对湿地松体胚的成熟影响很大,基本培养基以LP为最佳,ABA的最适浓度为10mg/L,PEG6000以添加25g/L、50g/L为最佳；在湿地松体胚的成熟培养过程中,PEG6000的浓度不能高于100 g/L；在湿地松体细胞胚的成熟培养基中,添加60g/L蔗糖最合适；湿地松体细胞胚的成熟培养基中添加0.5g/L或1g/L的活性炭最有利于体胚的成熟；培养基的状态对体胚的高频率诱导具有一定的影响,悬浮培养以100mL的三角瓶装30mL的培养液,摇床转速以130r/min为宜,液体转固体时吸取1mL培养液为宜；未建立成熟的体胚发生体系前,以采用固体培养为佳。最佳成熟培养基、激素及部分添加物组合为LP+10mg/L ABA+50g/L PEG6000+60g/L蔗糖+1g/L活性炭。     

不同类型供体细胞对牛体细胞重构胚发育能力的影响

通过显微操作去除体外成熟培养22￣23h卵母细胞的细胞核,并通过细胞质内注射法将供体细胞核移入去核卵母细胞内构建重组胚,重组胚经离子霉素(Ionomycin)激活5min后,再在6-DMAP内培养4h,将重组胚转CR1培养液中进行培养,24h和36h检查卵裂率,培养8d后统计囊胚发育率。比较了经血清饥饿和未饥饿牛成纤维细胞、颗粒细胞作为供体细胞构建的重组胚。研究发现:未饥饿组的牛成纤维细胞重组胚囊胚发育率(11.76%)显著高于血清饥饿组重组胚囊胚发育率(3.16%,p<0.05)。未饥饿组颗粒细胞囊胚发育率(14.94%)也显著高于血清饥饿组(7.14%,p<0.05)。经血清饥饿处理的颗粒细胞和成纤维细胞重组胚发育率差异不显著(p>0.05),未饥饿处理组两种细胞的重组胚发育率差异不显著。结果表明,用非饥饿处理牛成纤维细胞及颗粒细胞重组胚的发育率较高,牛颗粒细胞更有利于重新程序化。     

G418处理核供体细胞对猪克隆胚胎体外发育效率的影响

【目的】探索G418处理供体细胞对其核移植胚胎发育效率的影响。【方法】利用G418单独处理猪成体成纤维细胞6 d后,收集细胞,利用荧光定量PCR的方法检测处理前后细胞中抗氧化应激、细胞凋亡相关基因的表达水平,运用亚硫酸盐结合测序法分别检测基因组重复序列LINE-1、微卫星的DNA甲基化状态,以及其核移植胚胎体外发育的能力。【结果】经不同质量浓度G418处理的供体细胞的抗氧化应激酶相关基因及细胞凋亡基因表达发生显著变化(P0.05),但其DNA甲基化水平没有发生改变(P0.05);经G418处理的供体细胞的核移植胚胎的体外发育效率显著低于对照组(P0.05)。【结论】G418处理供体细胞可能对其克隆胚胎体外发育效率有抑制作用。     

3-acetylpyridine: effects in vitro related to teratogenic activity in chicken embryos

Production of skeletal muscle hypoplasia by 3-acetylpyridine and its complete reversal by nicotinamide in developing chicken embryos have been confirmed. Cultures of developing embryonic chicken muscle show degenerative effects produced by 3-acetylpyridine; these effects are reversed by nicotinamide. Cartilage production in cultured chondrogenic cells is potentiated by 3-acetylpyridine; this potentiation is completely reversed by nicotinamide. It is suggested that nicotinamide-or pyridine-nucleotide-dependent reactions influence normal differentiation of limb mesoderm cells by inhibiting chondrogenic-cell and potentiating muscle-cell expression or proliferation.     

紫斑牡丹‘书生捧墨’的体胚诱导与发生

为筛选适于紫斑牡丹体胚发生的诱导培养基，以‘书生捧墨’花后90 d 的种胚为外植体，采用L9(34)正交设计，研究了诱导培养基中基本培养基、蔗糖、BAP和NAA 4个因素及其组合对体胚发生的影响。结果表明:4个因素对体胚诱导率的影响均达到极显著水平，且以NAA对其影响最大，基本培养基和BAP次之，蔗糖的影响最小；4个因素的最佳组合为:改良MS+BAP 0.5 mg/L+NAA 0 mg/L+蔗糖100.0 g/L；经包含该组合的培养基培养70 d，体胚诱导率达63.3%，并获得了体胚再生植株。      

龙眼体胚发生中期发育同步化的初步调控

试验以龙眼"红核子"胚性愈伤组织为材料,对龙眼体细胞胚胎发生进行同步化调控,2,4-D浓度为0.07mg/L时获得了心形胚,2,4-D浓度为0.05mg/L和0.06mg/L时获得了鱼雷形胚,2,4-D浓度为0mg/L时获得了高度同步化的子叶形胚。研究表明,通过培养基中2,4-D浓度的微量调整,基本能够控制龙眼体胚发育中期心型、鱼雷形胚的发育同步化。