斑马鱼(Danio rerio)的资源管理：综述
斑马鱼胚胎发育中神经元迁移的调控
斑马鱼胚胎发育中神经元迁移的调控斑马鱼（Danio rerio）是一种被广泛用于生物学研究的水生动物，因其易于繁殖和培养，以及胚胎发育透明，方便研究和观察而备受关注。
在斑马鱼胚胎发育过程中，神经元的迁移是其中一个重要的过程，对于神经系统的形成与功能的发挥有着重要的影响。
因此，研究斑马鱼胚胎发育中神经元迁移的调控机制有着重要的科学意义。
神经元迁移的定义和重要性神经元迁移是指在神经系统发育中，神经元从胚胎前体向特定的位置前进而且改变形态的过程。
神经元迁移是神经系统正常发育过程中的一个关键环节。
在神经元迁移期间，神经元会通过定向移动和膜形态改变的方式进入其指定的位置，并与其他神经元组成神经电路。
这些神经电路是形成复杂神经网络的基础，控制着许多基本的生理行为和认知功能，如感觉、运动和行为学。
斑马鱼神经系统的发育斑马鱼胚胎发育的神经系统分为中央神经系统（CNS）和外周神经系统（PNS）两部分，CNS包括大脑和脊髓，而PNS包括不同形状和大小的神经节和神经丛。
斑马鱼胚胎发育的神经系统的结构和发育过程与哺乳动物相似，并且由于斑马鱼胚胎是透明的，这提供了研究神经系统发育和功能的难得机会。
神经元迁移的调控机制在斑马鱼神经系统的发育中，神经元迁移的调控是一个复杂、多因素的过程。
目前研究表明，神经元迁移涉及了多种信号通路和调节因子的参与，例如细胞间的信号分子和细胞自身的信号分子等。
在神经元迁移开始时，神经元需要定向向特定的位置前进，这个过程的调控是非常重要的。
许多分子参与了神经元定向移动和膜形态改变的过程。
其中一些分子是在神经元和周围细胞之间相互作用的信号分子，例如神经生长因子（NGF）和神经细胞粘附分子（NCAM）。
这些信号分子介导神经元与其周围环境的交流，确定神经元的运动方向和路径。
另一些重要的分子是细胞识别分子和结构蛋白，它们参与了神经元膜形态的改变和移动速率的控制。
但是，这些参与神经元迁移的分子的调控和相互作用模式仍然不完全了解。
斑马鱼卵黄原蛋白受体的功能分析
斑马鱼卵黄原蛋白受体的功能分析斑马鱼卵黄原蛋白受体的功能分析摘要:斑马鱼（Danio rerio）作为一种重要的实验动物模型，近年来在生命科学研究中越来越受到关注。
卵黄原蛋白是斑马鱼早期发育过程中的关键调控因子，起着重要的生物学功能。
本文通过系统性的功能分析，探讨了斑马鱼卵黄原蛋白受体在胚胎发育、胚胎毒理学等方面的功能机制，为进一步深入研究斑马鱼卵黄原蛋白受体的作用提供了理论依据。
关键词：斑马鱼，卵黄原蛋白受体，胚胎发育，胚胎毒理学，功能分析引言在生物体早期发育过程中，卵黄原蛋白扮演着极为重要的角色，其中一类卵黄原蛋白是通过卵黄原蛋白受体介导结合和内化起作用的。
斑马鱼作为一种优秀的实验动物模型，卵黄原蛋白受体在斑马鱼的胚胎发育中起着非常重要的作用。
本文将重点从斑马鱼卵黄原蛋白受体的功能出发，探讨其在胚胎发育和毒理学过程中的生物学作用和机制。
一、胚胎发育中的功能1. 体轴形成斑马鱼胚胎的体轴形成过程包括胚中胚层的形成和胚胎体轴的形成。
通过实验观察发现，卵黄原蛋白受体的存在能够影响斑马鱼的胚中胚层形成，从而影响体轴形成的过程。
2. 胚胎发育的调控卵黄原蛋白受体对斑马鱼胚胎发育中多个关键调控因子的表达进行了调控。
例如，在胚胎脊椎骨发育过程中，卵黄原蛋白受体能够调控促细胞分化和生长因子的表达，从而影响脊椎骨的发育。
3. 神经发育卵黄原蛋白受体在斑马鱼神经发育中发挥着重要作用，它通过调控神经营养因子的表达和神经元的生成，影响神经元的迁移和突触形成。
二、胚胎毒理学的功能1. 胚胎毒理学研究需求胚胎毒理学是一门研究外界物质对胚胎发育影响的学科。
斑马鱼由于其繁殖周期短、胚胎发育透明等特点，被广泛应用于胚胎毒理学研究中。
卵黄原蛋白受体能够与一些有机或无机化合物结合，从而影响斑马鱼的胚胎发育过程。
2. 卵黄原蛋白受体对毒物的敏感性卵黄原蛋白受体作为特异性受体，对毒物的敏感性较高，能够很好地反应斑马鱼胚胎对毒物的毒性效应。
斑马鱼模式生物简介
姓名：谭克强专业：生物技术学号：2009211803斑马鱼模式生物简介斑马鱼是在印度和巴基斯坦河里发现的一种鲤鱼。
成年3~4cm 长，有漂亮的花纹，群居生活。
在实验室里，斑马鱼广泛用于标准毒理学检验。
1996年，一系列论文报道斑马鱼胚胎发育的突变体的筛选及鉴定[1]，揭开了斑马鱼广泛应用于基因组功能分析、获得与脊椎动物发育和疾病相关的新基因研究的序幕。
斑马鱼(Danio rerio)作为水生脊椎动物的代表, 是现代遗传学、细胞生物学及发育生物学等研究的常用模式动物。
生物信息学预测认为, 斑马鱼基因组可能编码超过 400 种 miRNAs[2]。
通过构建不同发育阶段斑马鱼的小 RNA cDNA 文库发现, 斑马鱼miRNAs总数已经达到217个[2], 其中一些miRNAs在斑马鱼中的功能已经被解析。
miRNAs整体缺失对斑马鱼胚胎发育的影响研究结果表明, 缺失 miRNAs 的斑马鱼胚胎早期发育过程明显缓慢, 最初的 24 h 发育进程就被延迟了 3~4 h。
在原肠胚发育过程中, 突变胚胎不能进行正常的外包和内卷, 正常胚胎的索前板迁移发生在 80%外包时期, 而突变胚胎由于外包的推迟, 索前板迁移发生在 50%~60%外包时期。
此后, 由于体轴延伸减少导致胚胎缩短和脑部区域细胞的积累。
而在发育晚期, Dicer 突变胚胎后部卵黄延伸的范围也减少[3]。
Dicer 突变严重影响了神经胚形成。
由神经板发育成神经管的过程不能正常完成, 使神经管变成一个实心的棒状结构。
Dicer 突变胚胎脑内缺乏脑间隔而导致脑室数减少, 神经管腔的缺失和神经底板的减少表明脊髓发育也被干扰。
另外, 视网膜的发育也受到影响。
尽管神经系统发育畸形, 基因表达分析却发现, 神经管的前-后轴和背-腹轴图示均没有被完全破坏, 说明胚胎神经系统的图式形成和命运决定过程受到 miRNAs 的影响较少, 而脑的正常发育和神经细胞分化却需要 Dicer 酶的作用[3]。
斑马鱼综述写作5
Developmental Expression of the Nfe2-Related Factor (Nrf) Transcription Factor Family in the Zebrafish, Danio rerioLarissa M. Williams1,2, Alicia R. Timme-Laragy1, Jared V. Goldstone1, Andrew G. McArthur3, John J. Stegeman1, Roxanna M. Smolowitz4, Mark E. Hahn1*1 Biology Department, Woods Hole Oceanographic Institution, Woods Hole, Massachusetts, United States of America,2 Biology Department, Bates College, Lewiston, Maine, United States of America,3 Andrew McArthur Consulting, Hamilton, Ontario, Canada,4 Department of Biology and Marine Biology, Roger Williams University, Bristol, Rhode Island, United States of AmericaIntroductionThe nuclear factor-erythroid-2 (NFE2)-related factor (NRF) genes encode transcription factors in the Cap’n’Collar basic leucine zipper (CNC-bZIP) gene family. Members of this family include NFE2 [1,2], NRF1 (NFE2L1) [3,4], NRF2 (NFE2L2) [5], and NRF3 (NFE2L3) [6]. (For nomenclature conventions, see footnote 3 of ref [7]..) These transcription factors occur in vertebrates from fish [8] to mammals [9,10], and are known to regulate metabolic [11,12], cell cycle [12-14] and cytoprotective processes [9,13,15-18]. In mice, Nrf proteins have been shown to play important and often essential roles in development. Nfe2-null mice lack circulating platelets [19], the Nrf1 knockout is embryonic lethal [20], Nrf2-null mice are highly sensitive to carcinogens and oxidative stress [21], and the Nrf3knockout does not show abnormalities under normal conditions but is susceptible to lymphoma induced by exposure to carcinogens [22].Although studies in mice have provided important information about the function of Nrf genes, viviparous development makes it difficult to directly observe the role and effects of genes throughout the embryonic period.Zebrafish are a powerful model for studying molecular mechanisms of vertebrate development. Genes and signaling pathways [23,24] and molecular mechanisms of developmental toxicity [25-29] are highly conserved between fish and mammals. Furthermore, due to the whole genome duplication that occurred during early teleost radiation [30-33], zebrafish have duplicated copies (paralogs) that are co-orthologs of many single copy human genes [30,34]. Paralogs that have undergone subfunctionalization provide an opportunity for newmechanistic insights and thus new understanding about the functions of their single human counterpart [32,35]. In zebrafish, there are several paralogs within the nrf gene family, including nrf1a and nrf1b as well as nrf2a and nrf2b [7]. In total, zebrafish have six nrf family genes.In mammals, NFE2 forms a heterodimeric transcription factor with small MAF proteins, regulates expression of globin genes [2,36], and may regulate the oxidative stress response in mature erythrocytes [17]. In zebrafish, nfe2is a single copy gene [8]; its duplicate appears to have been lost [7]. In embryos nfe2 is highly expressed in the intermediate cell mass during erythroid differentiation [8]. Apart from in situ hybridization during development and protein characterization, the expression and role of nfe2in development are largely unexplored.NRF1 in mammals is expressed in many tissues [4]; it plays a role in regulating redox balance in the liver [37] and in regulating genes involved in development [20], oxidative stress [38,39], cytoskeletal organization [40], and the proteasome [41]. In response to UVB damage, mammalian NRF1 regulates nucleotide excision repair and glutathione homeostasis, and may act as a tumor suppressor [13]. The two zebrafish nrf1 paralogs, nrf1a and nrf1b, are syntenic with the duplicated hoxb clusters, similar to the human NRF1 that is near the human HOXB cluster [7]. Prior to the current study, the only information about expression of zebrafish nrf1had been from EST data [38] and a limited assessment by qualitative RT-PCR [7].NRF2 has a broad tissue distribution in mammals and acts as a pleiotropic transcription factor involved in regulating the susceptibility to disease, in mediating the response to xenobiotic exposure and oxidant stress, and in proteome maintenance [42]. In zebrafish, two paralogs (nrf2a and nrf2b) are broadly expressed both in the embryo and adult and apparently have undergone subfunctionalization [7]. The primary role of nrf2a seems to be in the regulation of cytoprotective genes upon oxidative stress [7,9]. Conversely, nrf2b is a negative regulator of embryonic gene expression both basally and under oxidative stress conditions [7].NRF3 has been found to be expressed at high levels in mammalian placenta and the B cell lineage and at lower levels in the heart, lung, kidney, and pancreas [6]. NRF3 may play a protective role in regulating genes responding to oxidative stress, as Nrf3-deficient mice treated with an oxidant suffer acute lung damage [43]. Zebrafish have one nrf3 ortholog, and like nfe2, the duplicated gene appears to have been lost [7]. The expression and function of nrf3in zebrafish was completely unexplored prior to this study.In this paper, we address fundamental aspects of the expression and role of six nrf genes during vertebrate development using the zebrafish as a model system. We show that the expression of nrf genes varies temporally throughout development and that nrf genes are induced following a pro-oxidant exposure and are potentially cross-regulated by gene family members Nrf2a and Nrf2b. We also identify a novel role for nfe2 in the cellular organization of epithelia in the pneumatic duct.Experimental ProceduresFish husbandryZebrafish of the Tupfel/Longfin mutation (TL) wild-type strain were used in all experiments. Adults were maintained and embryos were collected as previously described [44]. This study was carried out in strict accordance with the recommendations in the Guide for the Care and Use of Laboratory Animals of the National Institutes of Health. The protocol was approved by the Woods Hole Oceanographic Institution Animal Care and Use Committee (Animal Welfare Assurance Number A3630-01).Cloning and confirmation sequencing of nrf genesIn order to determine nrf cDNA sequences in the TL wild-type strain, and identify strain-specific polymorphisms, nrf cDNAs were amplified with primer pairs (Table 1), using Advantage cDNA Polymerase. The cycling conditions were [94°C, 1 min], [94°C, 30 sec; 68°C, 3 min] for 35 cycles, followed by 68°C for 3 minutes and 10 minutes at 15°C. The PCR products were purified with the GENECLEAN kit (Qbiogene, Quebec, Canada), ligated into pGemT Easy Vector System (Promega, Madison, WI) and constructs were sequenced. Prior to confirmation of in vitro expression, gene products from pGemT were subcloned into pcDNA3.1/Zeo (+) (Invitrogen, Carlsbad, CA). Sequences of clones were deposited into Genbank and have accession numbers JX867113-JX867116.Sampling, Chemical exposure, RNA extraction, and cDNA synthesisEmbryos for Developmental Series.As described in Timme-Laragy et al. [7], 4 pools of 30 staged embryos from a single clutch were reared at low density at 28.5°C in 0.3x Danieau’s and flash frozen in liquid nitrogen at 6, 12, 24, 48, 60, 72, 96, and 120 hpf and stored at -80°C. Hatched and unhatched embryos were collected separately at the 48 and 60 hpf time points. Unfertilized eggs, representing the 0 hpf time point, were manually stripped from 3 females, combined, and flash frozen as 3 separate biological replicates and stored at -80°C. Although it limited our ability to determine individual variation amongst embryos, pooling of embryos was needed to provide sufficient amounts of RNA for measuring expression of multiple genes.Chemical exposure of embryos to tBOOH.Embryos (3 pools of 30 staged embryos) were exposed to tert-butylhydroperoxide (tBOOH). Embryos (96 hpf) were exposed to 0, 0.5 or 0.75 mM tBOOH for 6 hours in 150x20mM glass petri dishes. Following tBOOH exposure, embryos were immediately placed in RNAlater and stored at -80°C.RNA extraction and cDNA synthesis.Total RNA was isolated from pooled embryo samples using RNA STAT-60 (AMS Biotechnology, Abingdon, UK) following the manufacturer’s protocol. cDNA was synthesized from 1 µg of total RNA using random hexamers and the Omniscript cDNA Synthesis Kit (Qiagen, Valencia, CA).Quantitative real-time RT-PCR gene expression profilingUsing the MyiQ Single-Color Real-Time PCR Detection System (Bio-Rad, Hercules, CA), QPCR was conducted with the iQ SYBR Green Supermix (Bio-Rad) on nrf genes. For each sample, duplicate reactions in separate wells were run containing 5 ng of cDNA. The PCR conditions were 95°C for 3.5 minutes followed by 35 cycles of 95°C for 15 seconds and 25 seconds at a gene specific temperature. Gene-specific primers and temperatures are listed in Table 1. Following each run, a melt curve was generated to ensure the amplification of single product. All primers were tested for amplification efficiency using a calibration dilution curve and slope calculation approach [45]. β-actin 1 (actb1) was chosen as a housekeeping gene due to its limited variation in expression with embryonic development and chemical exposure [46]. We confirmed the suitability of this housekeeping gene with the set of samples used in this study. Expression of genes wasanalyzed using the comparative ∆∆CT method [47].Statistical analysis of QPCR dataDevelopmental Series. Data were analyzed with MicrosoftExcel using the ∆∆CTmethod to calculate fold change. Valuesare presented as mean ± SEM, and N is defined as the numberof pools of embryos. Relative expression was normalized to the120 hpf value within a gene. As described in Timme-Laragy etal. [7], Statview for Windows (version 5.0.1; SAS Institute,Cary, NC) was used to determine differences in expression dueto hatching state. Data were log-normalized and six statisticaloutliers were removed from the nrf2b development series (onedata point from each of six time points: 0, 6, 24, 48(unhatched), 48 (hatched), and 96 hpf). If an ANOVA yieldedsignificance (p< 0.05), Fisher's protected least significantdifferences test was used as a post hoc test with Bonferronicorrection.Chemical Exposure and Nrf2a and Nrf2b crosstalk. Datawere analyzed with Statview for Windows (version 5.0.1; SASInstitute, Cary, NC) and presented as mean ± SEM where N isdefined as the number of pools of embryos. Significance inANOVA was determined with a p≤ 0.05. When significancewas yielded, Fisher’s protected least-significant differences testTable 1. Primers used for gene amplification, cloning, and qPCR of nrf genes.Gene Primer sequences (5’-3’)Amplicon size T (°C)RefR-GGTGAACTATCACTTTAATCAAACATnrf1a F-CAGTTCGCACGCCCTTATTTACTGAC237668-R-GCTGATGGACTTAACAGCAGACAGnrf1b F-CGTAACCTAATTTGGTTTGACG288568-R-GTCCTCCTTGACTTCCCATATCnrf3F-AAATTGAGCAGTTGCTCCCCTCC234668-R-CTCACCTCAAAGATAAAACTCACCnfe2-qPCR F-CAGAGTTTGAGGAACCCAATGAG12757-R-CACAAGTGGCTGGAATGGATTCnrf1a-qPCR F-CCAGAGTTGACAGGTCCTGG15864-R-CATAACCTGTGATTCCATGATAGACnrf1b-qPCR F-GCAGGACATGGAGGTGAACAATACG12066-R-GGATCGTGGGAGCCCAAAATTTCCnrf2a-qPCR F-GAGCGGGAGAAATCACACAGAATG8265[7] R-CAGGAGCTGCATGCACTCATCGnrf2b-qPCR F-GGCAGAGGGAGGAGGAGACCAT26968[7] R-AAACAGCAGGGCAGACAACAAGGnrf3-qPCR F-GCATGAGGATTTAGTGGTTAGTGG10864-R-GGAGTCAAAATCATCAAAGTCAGactb1-qPCR F-CAACAGAGAGAAGATGACACAGATCA14065[87] R-GTCACACCATCACCAGAGTCCATCACPrimer sequences are written 5’-3’ and amplicon size and melting temperature (T) are shown. For primers used previously, a reference is cited. Accession numbers for sequences newly reported here are: nrf1a (JX867114), nrf1b (JX867116), nrf3 (JX867113), nfe2 (JX867115).doi: 10.1371/journal.pone.0079574.t001was used as a post hoc test with Bonferroni correction. Relative expression was normalized to the water control for tBOOH. For Nrf2a and Nrf2b morpholino data, relative expression of each gene was normalized to the water control with control morpholino.Microarray gene expression profilingZebrafish embryo rearing, sampling, and microarray analysis were performed as previously reported [29]. Briefly, four replicate pools of 100 embryos were examined at 3, 6, 12, 24, 36, and 48 hpf. Agilent 4 × 44k DNA gene expression microarrays (part #GPL11077, WHOI Zebrafish 44k v1.0 custom-commercial Chemical Defensome array, Agilent Technologies, Santa Clara, CA), which included custom probes to ensure adequate coverage of the chemical defensome [29], were used to examine gene expression levels. Single color microarray data (Cy3) was normalized using the non-linear scaling method of Schadt et al. [48], where saturated probes and probes not above background in all replicated were removed. Because standard ANOVA is not appropriate for these data due to auto-correlation between time points, we used Bayesian Estimation of Temporal Regulation (BETR; [49]) to analyze the developmental time series. Normalized Cy3 values for each probe were log transformed, median-centered, and analyzed using BETR relative to 3 hpf. Normalized average Cy3 signals were compared for genes of interest among time points. All probes reported here had significant differential expression among sampling times. The microarray data have been deposited into the Gene Expression Omnibus (GEO) database with NCBI GEO accession number GSE24840.In situ hybridizationIn situ hybridization of nrf3 on whole embryos was performed using established procedures [50,51]. Sense and anti-sense nrf3 RNA probes were created by restriction digest of the nrf3 construct (in pGemT) with XhoI, yielding a 1080 bp sense probe and 1328 bp anti-sense probe. The nrf3 probe covers a 1240-bp region of the 3’ end of the cDNA and includes 87 base pairs of the 3’ UTR. In designing the probe, we carefully considered possible sequence identity with other nrf transcripts. Nucleotide sequence identities between nrf3and other nrf transcripts in the region covered by the probe were as follows: 45% with nfe2, 47% with nrf1a, 51% with nrf1b, 49% with nrf2a, and 45% with nrf2b. The overlap between the nrf3 probe and other nrf transcripts was typically interspersed throughout the sequence with no long runs of overlap (mainly, 4-5 nt at a time). We also conducted a Blast search using the nrf3 probe against the zebrafish genome; no significant overlap was found other than the nrf3 gene itself. Probes were labeled with digoxigenin-UTP by in vitro transcription (Roche San Francisco, CA) with either T7 (sense) or SP6 (anti-sense) polymerase. Pre-hybridization was carried out for 2 hours at 65°C. Hybridization was carried out overnight with 300 ng of probe per pool of seven embryos at a concentration of approximately 1.5 µg/mL. Embryos were blocked for 4 hours at room temperature and incubated with anti-digoxigenin alkaline phosphatase conjugated antibody (Roche, San Francisco, CA)at 4°C overnight. Embryos were imaged by light microscopy (Axio Observer.A1, Carl Zeiss, Oberkochen, Germany).Morpholino oligonucleotidesMorpholino antisense oligonucleotides (MO) were designed to block initiation of translation of zebrafish nfe2, nrf1a, nrf1b and nrf3 and obtained from Gene Tools, LLC (Philomath, OR). The first nfe2MO (AGTTCCTGCCAGGCCAAGTCCATCT) complements two residues of the 5’UTR, the start codon (underlined), and 20 residues of the coding region. A second, non-overlapping nfe2MO (AACGATGTGTCCGTAATCCAGTGAC) complements 25 residues of the 5’UTR and targets a region whose 3’ end is 15 residues upstream of the sequence targeted by the first nfe2 morpholino. The nrf1a MO (ATGGCCCAAACCATCACCGGCAGCA) complements 11 residues of the 5’UTR, the start codon (underlined), and 11 residues of the coding region. The nrf1b MO (AATCACGCAAACAAACGTCAAACCA) complements 25 residues of the 5’UTR and its 3’ end is located 34 residues upstream of the start codon. The nrf3MO (TTTTAACCTCAGGAGGCTTAAACGA) complements 25 of the 5’UTR and its 3’ end is located 14 residues upstream of the start codon. The standard control-MO from Gene Tools was also used (CCTCTTACCTCAGTTACAATTTATA). All MO were tagged at the 3’ end with fluorescein in order to detect successful injection and incorporation of MO into embryos. MOs targeting nrf2a and nrf2b are previously described [7]. Confirmation of MO translation inhibition by in vitro protein synthesisTo confirm the efficacy of morpholinos to inhibit translation of Nrf proteins, the TnT T7 Quick Coupled Reticulocyte Lysate System (Promega, Madison, WI) was used to synthesize [35S]methionine-labeled zebrafish NF-E2, Nrf1a, Nrf1b and Nrf3 protein as per manufacturer’s protocols and as described previously [52]. Briefly, the TnT reaction included 1 μl of [35S]methionine (> 1000 Ci/mmol at 10 mCi/ml) and 2 μl of the respective nrf cDNA in pcDNA3.1/Zeo (+) (0.5 μg/μl) in a final volume of 25 μl with H2O. For reactions containing MOs, 0.5 μl of a 25 μM stock of the appropriate MO was added, resulting in a final concentration of 500 nM. Labeled protein products were resolved by SDS-PAGE, dried on Whatman filter paper, and visualized on film. To quantify the reduction in protein products, gel fragments containing a single protein band were excised and placed into 7 mL glass scintillation vials containing 5mL ScintiVerse II cocktail (Fisher Scientific, Pittsburg, PA). Counts were carried out for 5 minutes on a Beckman Coulter LS6500 Multi-purpose Scintillation counter (Brea, CA) and background was subtracted.Microinjection of zebrafish embryos with morpholinos Zebrafish embryos at the two- to four-cell stage were injected with 2.5-5 nL of a 0.1 mM solution of MO targeting nfe2, nrf1a, nrf1b, nrf2a, nrf2b, nrf3, or a control-MO, using a Narishige IM-200 microinjector (Tokyo, Japan). Injection volumes were calibrated as described previously [52]. The injections resulted in delivery of MO amounts ranging from 2-4 ng. At six to ninehpf, embryos were sorted and screened for successful fertilization and fluorescence. When no developmental phenotypes were observed, the concentration of the morpholino stock was increased to 0.15 mM, keeping the injection volume constant. MO-injected embryos were held in 0.3x Danieu’s solution. At 4 dpf, swim bladder inflation was scored as described earlier [53].Histological analysis of the swim bladderFour-day-old larvae from embryos injected with control morpholino (swim bladder inflated) or nfe2morpholino (swim bladder not inflated) were fixed in 4% formaldehyde in 1x phosphate buffer, dehydrated in ethanol and stored in 75% ethanol until embedding. Larvae were sent to Environmental Pathology Laboratories (Sterling, VA) where they were embedded laterally into either Technovit 7100 (Heraeus Kulzer, Hanau, Germany) or paraffin. Sagittal sections were made serially every 2 µm. Sections were mounted on superfrost glass slides and stained with hematoxylin and eosin. Histopathology was conducted by an Olympus BX 40 with an Olympus DP25 camera system.Analysis of blood flowTransgenic gata1dsRED embryos [54] (15 individuals per treatment) were injected with control or nfe2-MOs at one- to four-cell stages. As an index of blood flow, the number of red blood cells passing through the mesencephalic vein (MsV) per 15 s was determined by orienting 48 hpf zebrafish embryos laterally as described by Kubota et al. [55]. Blood flow was recorded with an AxioCam MRc5 camera on a Zeiss Axiovert 200 inverted microscope and analysis (counting of red blood cells) was performed using ImageJ (/ij/).Analysis of otic vesiclesAB embryos were injected at one- to four-cell stages with control-MO or nfe2-MO-1. At 30, 48 and 72 hpf, otic vesicles were imaged with an AxioCam MRc5 camera using a Zeiss Axiovert 200 inverted microscope.In silico Promoter Analysis and motif searchesTo determine whether Nrf2a or Nrf2b are directly involved in the transcriptional regulation of other nrf family members, in silico promoter analysis was carried out for AREs and XREs using a fuzzy search algorithm, fuzznuc [56]. 10,000 base pairs upstream of the start site and the entire length of the gene were examined for putative zebrafish AREs (TGA(G/C)nnnTC [57]) and XREs (KNGCGTG [58]). Additional searches were carried out for Nfe2 binding sites [59] determined from ChIP-Seq studies. The zebrafish genome was searched for Nfe2 binding sites in the 10 kb upstream of the start codon of all known genes using a position-specific scoring matrix (PSSM) based on the empirical ChIP-Seq results of Wang et al [59] and the FIMO algorithm of the MEME/MAST package [60](p < 0.0001), with correction for background nucleotide frequencies of complete nuclear genome sequences.ResultsDevelopmental expression profilingTo begin to understand the role of the nrf genes during development, their expression was profiled from 0 to 120 hpf using qRT-PCR (Figure 1). Because the data were normalized within each gene, expression values can be compared at different times for each gene but not across genes. nfe2 transcripts were maternally deposited, but levels fell substantially at 6 hpf and did not increase again until 48 hpf. Expression remained relatively constant from 48 to 60 hpf and then decreased steadily until 120 hpf. Like nfe2, nrf3was maternally deposited and decreased by 6 hpf, but transcript levels increased again at 12 hpf. Expression decreased between 12 and 48 hpf, but then remained steady. nrf1a was expressed from 0-24 hpf but levels were reduced from 48-72 hpf, after which expression increased at 96 and 120 hpf. Conversely, nrf1b expression peaked between 12 and 24 hpf and then declined and remained relatively low from 48 to 120 hpf. Hatching did not have a significant effect on expression of nfe2, nrf3, nrf1a or nrf1b. For comparison, Figure 1 also includes data for expression of nfr2a and nrf2b, originally presented (in a slightly different format) by Timme-Laragy et al.[7]. These results showed that nrf2a expression increased steadily during development. The expression of nrf2b also increased through 48 hpf, but there was a significant effect of hatching on nrf2b expression, when expression decreased in hatched embryos compared to unhatched embryos of the same developmental age.Since molecule numbers were not available from the qRT-PCR data and data were normalized within each gene, a microarray was used to determine the expression of nrf genes relative to each other during the first 48 hours of development (Figure 2). Microarray and QPCR analyses were performed on samples obtained from independent experiments. The microarray data did not include a 0 hpf time point, so maternal transcript levels were not determined with this method. Gene expression profiles determined by QPCR and microarray for nfe2, nrf1a, nrf2a, and nrf2b displayed similar developmental patterns. The developmental patterns of nrf3expression measured by qPCR and microarray were similar, except that the pattern at 6 and 12 hpf was reversed. For nrf1b, the peak at 12 hpf measured by QPCR was not evident in the microarray data.Normalized Cy3 levels for nfe2, nrf1a, nrf1b, nrf2a, and nrf2b ranged from 100 to 1,800, whereas values for nrf3 ranged from 80,000 at 3 hpf to approximately 20,000 at 48 hpf (Figure 2). The nrf gene with lowest expression was nrf1a followed by nrf1b. Their average expression values were about half that of nfe2. The expression of nrf2a was up to10-fold greater than that of all other nrf genes except nrf3.Localization of nrf3 expression with ISHWe used in situ hybridization to examine the spatial expression of nrf3, the nrf gene most highly expressed during development; analysis was carried out in 12-hour intervals from 24 to 72 hpf. In situ gene hybridization results for nfe2, nrf1a, and nrf2a have been previously reported [8,61,62]. The mostprominent expression of nrf3 occurred at 24 and 36 hpf;expression was detected in the fore-, mid- and hindbrain as well as in the pectoral fin buds (Figures 3A, 3B, 3D, 3E). The high degree of staining seen is consistent with the relatively high level of nrf3 expression seen in the array and qRT-PCR data. At 48 hpf and 60 hpf, expression still was seen in the pectoral fin buds, and was less prominent in the forebrain andmore concentrated in both the mid- and hindbrain (Figures 3G,Figure 1. The expression of the nrf gene family during development as measured by quantitative real-time PCR. Black lines indicate expression pre-hatch and grey lines indicate expression post-hatch. Values were normalized to the 120 hpf time-point and β-actin 1 (actb1) was used as the housekeeping gene. Data are presented as the mean ± S.E.M. (error bars), and N = 4 pools of 30 embryos. Differences in expression between hatching state were assessed using ANOVA followed by Fisher's PLSD (*, p ≤0.05). Data for expression of nrf2a and nrf2b are from Timme-Laragy et al. [7] and are used with permission of the American Society for Biochemistry and Molecular Biology. Previously these data were presented as molecule number. Here we have normalized the data to β-actin 1 expression and to the 120 hpf time point and re-plotted for comparison to the new data on expression of other nrf family genes. Dashed lines indicate the value of 1.0 from the 120 hpf time point.doi: 10.1371/journal.pone.0079574.g0013H, 3J, 3K). Expression was also seen in the branchiogenic primordia. Expression at 72 hpf was almost exclusively in the mid/hindbrain junction (Figure 3M, 3N). There was no signal in sense controls at any time point assayed (Figures 3C, 3F, 3I,3L, 3O).Effect of nrf gene knockdown on developmentOne approach to identify the importance of nrf genes in developmental processes is to knock down their expression in the embryo. The in vitro translation-blocking efficiency of nrf morpholinos ranged from 65 to 68%, with the exception of morpholino 2 for nfe2 (22%; Figure 4). To determine whether any morphological abnormalities were associated with the reduction of Nrf proteins, embryos were injected with morpholinos and observed every 12 hours until 120 hpf. No gross morphological abnormalities were observed for control MO, nrf1a -MO, nrf1b -MO, a combination of nrf1a -MO and nrf1b -MO, or nrf3-MO. Additional experiments were conducted using higher concentration of morpholino (0.15 mM). However,at this concentration off-target effects were observed. Off-targeteffects observed in more than 50% of the MO-injected embryos included hatching gland deformities, pericardial edema, and bent tail post-hatching.In contrast to the lack of specific phenotypes in embryos injected with MOs targeting nrf1a, nrf1b , or nrf3, injection of an nfe2-MO (0.1 mM) resulted in the failure of the swim bladder to inflate at 96 hpf in 95% of the injected embryos (Figure 5B versus 5A) (Table 2). The specificity of this effect was confirmed using a second, non-overlapping nfe2 MO that also targeted the region surrounding the translation start site (Table 2).Histological analysis of larvae treated with control MO showed the normal pneumatic duct connection between the esophagus and the swim bladder (Figure 5C,D). Epithelial cells along the duct were orderly and cell morphology progressed from columnar at the entrance to the duct to cuboidal to squamous at the entrance to the swim bladder. In larvae in which Nfe2 had been knocked down, the pneumatic duct connection was intact but the number of epithelial cells and the regular progression from columnar to squamous was disrupted (Figure 5E). In the most severe example, the epithelialcellsFigure 2. The expression of the nrf gene family during development as measured by microarray analysis. Data are presented as normalized Cy3 levels, with average values for nfe2, nrf1a , nrf1b , nrf2a , and nrf2b displayed on the primary (left) y-axis. Values for nrf3 are displayed on the secondary (right) y-axis. Each time point represents the average normalized value ±SD of 4 replicates, each from 100 embryos. The Agilent probes from which these data were obtained are: nfe2 (A_15_P109440), nrf1a (A_15_P110831), nrf1b (A_15_P116909), nrf2a (CUST_121_PI358351581), nrf2b (CUST_122_PI358351581), nrf3(A_15_P116878). An additional probe for nrf2a (A_15_P109504) was present on the array and produced results very similar to those shown here for CUST_121_PI358351581. The microarray data have been deposited with NCBI GEO accession number GSE24840.doi: 10.1371/journal.pone.0079574.g002。
斑马鱼模式生物简介
斑马鱼模式生物简介斑马鱼（Danio rerio）是一种小型的鱼类，也是一种非常受欢迎的淡水观赏鱼，它是孔雀鱼科中的一员，是印度东北部和孟加拉国的淡水鱼类。
斑马鱼模式生物是国际上被广泛应用于基因工程和生物医学研究的模式生物之一，也可以作为重要的药物筛选平台和疾病模型使用。
那么，接下来就来逐一介绍斑马鱼的分类、形态特征、生活习性、模式生物优势及应用等方面的知识。
一、斑马鱼的分类斑马鱼属于脊索动物门、脊椎动物亚门、圆口纲、鲤形目、鲤科、孔雀鱼属的一种淡水鱼类，学名为Danio rerio。
斑马鱼还有各种不同的变种和突变体，例如白斑马鱼、红斑马鱼、紫斑马鱼、金斑马鱼等。
二、斑马鱼的形态特征1.外形特征斑马鱼身体呈纺锤形，通常长度为2.5-4厘米，雄性略大于雌性。
头部和尾部比身体稍大，呈楔形，头部较尖，口小，下颌略突出。
各种颜色和花纹的斑马鱼几乎可以说是身体通体条纹，由水平条纹串联俩个腹鳍之间的细密黑色点状线条逐渐变浓变宽而形成，其中背部、腹部及尾鳍的条纹分化明显。
因此，美国生物技术研究所把斑马鱼称为"斑马状鱼"（Zebra fish）。
2.内部结构斑马鱼的内部结构和人类相似，有头、躯干、尾等结构。
头部包括大脑、眼、鼻、口、耳等器官，身体包括骨架、肌肉、消化系统、心脏、循环系统、呼吸系统、泌尿系统等器官。
三、斑马鱼的生活习性1.水温适应性斑马鱼适应范围广，可在25摄氏度以下的水温生存，适应性强，而50摄氏度以上的水温则容易引起死亡。
通常在水温20度-28度之间生存。
2.繁殖生殖斑马鱼在2-3个月龄时性成熟，通常在水中产卵。
雌鱼体内产生卵子时，会有告诉性色素出现，这些色素可以帮助斑马鱼有效地控制自己和同伴的繁殖。
雄鱼会在巢穴内产生精子，和雌鱼配对后可以产生400-500颗卵。
在适当的条件下斑马鱼卵可以在69h后孵化。
3.食性习性斑马鱼性质温和，不挑食，食性杂，吃小型昆虫、蚯蚓、水虫以及水草、藻类等，在食物充足的情况下生长迅速，但在食物短缺时可以进入休眠状态。
(完整版)斑马鱼动物模型的应用介绍
斑马鱼动物模型的应用斑马鱼（Danio rerio）属于辐鳍亚纲（Actinopterygii）鲤科（Cyprinidae）短担尼鱼属（Danio）的一种硬骨鱼，原产于南亚，是一种常见的热带观赏鱼，因其体侧具有斑马一样暗蓝与银色相间的纹条而得名。
斑马鱼个体小，易于饲养，成体长4-5cm，雄鱼体修长，雌鱼体肥大。
可在有限空间里养殖相当大的群体，可满足样本需求量大的研究。
斑马鱼发育迅速，在28.5℃培养条件下受精后约40min完成第一次有丝分裂，之后大约每隔15min分裂一次，24h后主要器官原基形成，相当于28d的人类胚胎，幼鱼孵出后约3个月达到性成熟。
雌雄鱼通过调控光周期控制14:10（光照:黑暗）产卵时间，成熟鱼每周可产卵一次，一尾雌鱼每次可产卵100-300枚。
胚胎体外受精，体外发育，胚体透明，易于观察。
受精卵直径约1mm，易于进行显微注射和细胞移植等操作。
一、斑马鱼的品系经过30多年的研究应用和系统发展，已有约20个斑马鱼品系，斑马鱼基因数据库-ZFIN （http://zfin/org）里有相关的资料可供查询和下载。
目前研究中常用的斑马鱼野生型品系主要为AB 品系、Tuebingen（Tu）品系、WIK 品系，斑马鱼基因组计划所用品系是Tu。
AB 品系是实验室常用的斑马鱼品系，由单倍体细胞经早期加压法获得。
Tu品系斑马鱼具有胚胎致死突变基因，用于基因组测序前敲除该致死突变基因。
WIK品系较Tu品系具有更多的形态多样性。
此外，还保存有3000多个突变品系和100多个转基因品系。
这些品系资源对于利用斑马鱼开展各种科学研究起着很大的推动作用。
二、斑马鱼突变品系的筛选斑马鱼突变的方法主要有三种：已基亚硝脲（ENU）化学诱导、γ或χ射线照射和插入诱变。
ENU是一种DNA烃基化试剂，在生殖细胞减数分裂前诱导碱基对的替换，诱导产生的突变率为0.1%-0.2%，涉及单个基因的突变。
射线照射导致染色体大片段的缺失或染色体重排，产生突变率达1%。
华中农大：斑马鱼养殖综述
斑马鱼(Danio rerio)养殖综述Christian Lawrence布莱根妇女医院卡帕研究实验室马萨诸塞州美国收稿日期：2007年7月16日；改修日期：2007年8月24日；接受日期：2007年8月25日。
摘要：斑马鱼(Danio rerio)是脊椎动物研究的模式生物。
斑马鱼的优良性状有利于研究人类疾病的发生与传播。
较强的繁殖力、短小的体型、繁殖周期短、胚胎早期光学透明性强等特性使得斑马鱼成为许多研究者研究其他项目青睐的对象，包括研究动物行为、鱼类生理机能和水产养殖毒性试验。
尽管如此，科学严谨的斑马鱼养殖技术还是很不发达。
尽管斑马鱼有很大的需求量，包括直接或者间接养殖。
斑马鱼的养殖缺乏应用养殖标准。
本文试图将已有的关于斑马鱼生物学和养殖的科学信息进行综述，可以用来提高这一重要的模型动物使用检索的效率。
本研究还强调了该领域还需要进一步研究的内容。
关键词：斑马鱼；Danio rerio；饲养；水产养殖；管理目录：1、引言 (2)2、斑马鱼自然生活史 (3)2.1、分布和栖息地 (3)2.2、生殖和行为 (3)2.3、寿命 (5)2.4、食物 (5)3、斑马鱼养殖 (5)3.1、水化学参数 (5)3.1.1、温度 (6)3.1.1、pH (6)3.1.3、硬度 (7)3.1.4、盐度 (7)3.1.5、溶解氧 (8)3.1.6、含氮废弃物 (8)4、营养、食物、投喂方式 (8)4.1、营养需求 (8)4.2、食物 (9)4.3、投喂 (11)5、繁殖和育种技术 (11)5.1、繁殖 (11)5.2、育种技术 (12)5.3、产卵效率 (13)6、幼鱼饲养 (13)6.1、幼鱼生物学 (13)6.2、食物和营养 (13)6.3、水质 (14)6.4、生长率和存活率 (15)7、成鱼培育 (15)7.1、放养密度 (15)7.2、遗传育种 (16)8、结语 (16)鸣谢 (16)参考文献 (16)1、引言在过去的二十年间，斑马鱼(Danio rerio)广泛作为遗传学和发育学研究的优良模式生物(Fishman,2001)，最近斑马鱼还应用于的人类疾病和治疗药物筛选的研究当中(Penberthy et al.,2002;Sumanasa and Lin, 2004)。
斑马鱼研究计划与相关资源简介_肖安
HEREDITAS (Beijing)2013年4月,35(4):547―548ISSN 资料库网络出版时间:2013-1-309:54:36URL:/kcms/detail/11.1913.R.20130130.0954.001.htmlDOI:10.3724/SP.J.1005.2013.00547斑马鱼研究计划与相关资源简介肖安,张雨田,张博北京大学生命科学学院,细胞增殖与分化教育部重点实验室,北京100871斑马鱼(Danio rerio )是目前遗传发育研究领域最为活跃的模式脊椎动物之一,有多个大型研究计划正在进行中。
本文选择其中的一部分进行简要介绍。
1斑马鱼基因组测序计划斑马鱼基因组测序计划始于2001年,最初由英国Sanger 研究所主持,现在转由参考基因组协会(Genome Reference Consortium,GRC)主导,跟人类、小鼠两个基因组计划一起成为GRC 的三个主要目标。
测序数据来自基于克隆图谱测序法和全基因组鸟枪法的整合结果。
随着测序进度的发展,主持单位会定期更新斑马鱼基因组的拼装,最新版本为2010年7月发布的Zv9。
目前斑马鱼基因组计划即将完成,已进入缺口(gap)修补和未定位片段定位的阶段,GRC 计划于2013年晚期发布新的拼装版本GRCz10。
在GRC 的计划页面上可以看到不断更新的缺口修补进度和其它信息。
斑马鱼基因组注释计划主要分为自动注释和人工注释两部分,分别发布于Ensembl 和Vega 数据库,此外,还有相当多方面的注释内容由UCSC Genome Browser 首先提供,但各个数据库间均相互共享数据。
2斑马鱼突变计划及相关的数据库(1)ZMP(Zebrafish Mutation Project)计划：原称Zebrafish mutation resource 计划,是由Sanger 研究所主持的一项基于TILLING(Targeting Induced Local Lesions IN Genomes)技术的斑马鱼突变筛选计划。
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斑马鱼（Danio rerio）的资源管理：综述Christian Lawrence摘要斑马鱼最近成为了一个卓越的生物医学研究的模型的脊椎动物。
它作为人类疾病和发展的模型，这个同样令人喜欢的特征为它受欢迎做出了贡献；即繁殖力高，体积小，迅速的繁衍周期，在早期胚胎发育早期的光学透明性，也有许多其他学科的研究者长期致力于它的研究，包括动物行为，鱼类生理，水产毒理学。
尽管如此，严谨的饲养斑马鱼的技术还是不够发达。
虽然斑马鱼有一个相当大的身体，都和畜牧业有直接或间接的关系，这个信息有许多不同的来源，而且它很少被应用到发展中国家的标准协议。
这项综述是尝试把可利用的与斑马鱼生物学和文化相关的科学资料整合到一个这项领域的概述，可以用于研究中的这个重要的动物模型的使用效率的改善。
这个综述还强调了在那些领域需要做进一步的研究。
目录1.简介2.斑马鱼的自然历史2.1 喜好的栖息地及分布2.2 繁殖和行为2.3 寿命2.4 食性3.斑马鱼文化3.1 水化学3.1.1 温度3.1.2 PH值3.1.3 硬度3.1.4 盐度3.1.5 溶氧3.1.6 含氮废物4.营养，食性和饲养方法4.1 营养需求4.2 食性4.3 饲养5.繁殖和养殖技术5.1 繁殖5.2 养殖技术5.3 产卵率6.幼体培育6.1 幼体生物学6.2 食性和营养6.3 水质6.4 生长率和存活率7.成体培育7.1 保持密度7.2 遗传育种计划8.总结致谢参考文献1.简介在过去的二十年里，斑马鱼已成为一个研究遗传学和发展的重要的脊椎动物模型，近来，也包括人类疾病和筛查治疗药物。
大量的有利的性质，包括它体积小，快速的发展和繁殖速度，早期发展过程中的光学透明性，比较容易的遗传选育和与人类相似的遗传特点，而且它将很有可能在其他领域的研究中刺激经济的增长，特别是它的基因组草案的进一步完善和令人振奋的用来做扩展研究的可利用的工具和方法。
鉴于斑马鱼作为一个相当重要的实验模型，伴随着的是它大量使用和相关培育设施的建立和维修的大额的经济耗费，在一定程度令人惊讶的是它的畜牧业不发达。
当与其他的养殖鱼类相比，像罗非鱼，斑点叉尾鮰和鲤鱼，公开的饲养标准完全不符合要求。
在有关养殖技术的信息交流中，众多的研究机构之间的房屋斑马鱼养殖技术已经基本不存在，和任何已经取得的进展都非常孤立，没有同行业内人士的利益评价。
这种情况的一个不幸结果是，许多斑马鱼的设施可能运作在一个次级水平，当调查人员初次接触这个模型，在为新建立的养殖设施制定一个标准的作业程序，没有科学严谨的资源可以咨询时，这个情况只会进一步加剧。
尽管公布的饲养标准缺乏，斑马鱼任然是一种非常好的研究动物。
很久以前，它作为一种发展模型出现，斑马鱼是研究各种鱼类各方面生物学的材料。
多年来，研究人员利用这些具体实际的优势，现在用斑马鱼的生物学研究团体来研究鱼类的繁殖,变浅，水生毒理学，渗透调节和嗅觉。
近来，一些团体都发表了有关斑马鱼的自然历史，营养和繁殖行为研究文章。
这研究历史可能帮助斑马鱼研究者拥有一个丰富资源的生物学研究机构，那个可以直接或间接地应用于畜牧业。
然而，当这些信息被发布后，不同的是，他发表在新闻期刊上后不会被斑马鱼研究团体的人们来做一般的咨询。
1.斑马鱼的自然历史一个对斑马鱼的自然环境更全面的了解，包括喜好的栖息地，繁殖行为和食性，对完善饲养标准和优化在生物医学和遗传行为方面广泛使用的研究都是必须的。
然而，发表的有关斑马鱼自然历史的信息是有限的，和那些落后于为这个物种的大量遗传和发育有利数据。
实际上，几乎没有可利用的有关斑马鱼生态和行为的资料被用来畜牧业的发展，但是这种趋势必将改变普遍的继续增长的模式，并且应用到不同领域的研究，比如说遗传行为。
这些信息对畜牧业的重要性，被来自少数领域研究数据所证实，这些研究已经被多次引用或者出现在本文的更多部分。
下面是对这些可利用数据的一个概要的总结，包括可以被用来改善斑马鱼饲养研究设施的更长远工作的建议。
一个斑马鱼更充分的行为和生态的治疗将由斯彭斯等人给出。
2.1 喜好的栖息地及分布斑马鱼原产于南亚，主要分布在印度，孟加拉国，尼泊尔，缅甸和巴基斯坦的部分地区。
这个地理区域里有季风气候，明显的雨季和旱季对它的栖息地分布有明显的影响，包括水化学性质和丰富的资源。
斑马鱼已经被报导出现在这个区域里的多种多样的栖息地类型，包括灌溉的沟渠和稻田，人造的池塘，河流的上游，甚至是湍急的溪流。
然而，在孟加拉国和印度进行的两次调查结果显示，把这两次结果合并到一起，提供了一个至今对这个物种喜好栖息地最全面的描述。
麦克卢尔等人进行了六次采样调查，包括位于西孟加拉邦和北方邦的印度各邦的恒河流域，在这三个地方发现了斑马鱼，其中包括稻田和两个山脚溪流的静水流。
在孟加拉国，在恒河和雅努藏布江流域的23个地方的调查中发现斑马鱼在这些位置9。
在印度的研究调查中，斑马鱼仅在静水或缓慢流动的水域中发现，包括浅水池塘，其中还有一部分用来种水稻。
有趣的是，斑马鱼均与这些地方的大部分或全部水生植被有联系。
值得注意的是，在野外或以实验室为基础对野生或家养的斑马鱼的行为研究中，它们更倾向与在与其水生植被有关系的地方产卵。
这些调查还描述了更多的发现斑马鱼的地方的水的物理化学性质，包括PH值，温度，盐度，透明度，深度和百分之冠（见表一）。
总之，这些观察表明，斑马鱼似乎更倾向于静止或者缓慢地移动，透明度高的弱碱性的水体。
这些数据是有用的，至少它们可能直接用于实验室斑马鱼的水化学的参数的优化设计，当鱼被放流在与自然相适应的水环境中时，它的人工饲养生产力是最高的。
今后的实地研究应该更充分地调查在夏季的情况，在一个更宽广的栖息地类型，以确定斑马鱼在自然界中全部的喜好栖息地，从而使之更好的配合实验室的水化学参数成为可能。
2.2 繁殖和行为由于斑马鱼的研究在很大程度上是以生产大量的育苗为前提的，野生动物的生物繁殖和行为的信息与畜牧业明显相关。
值得注意的是，很少有人知道这方面的物种。
许多可利用的数据主要来源于动物调查期间收集者的意见，尽管许多更加详细的说明已经被报导，尤其是最近的。
迄今为止收集的证据表明，斑马鱼出现缓慢流动水体的小沙洲中或者漫滩水体的静水中。
斑马鱼是体外繁殖，小群体的批次产卵繁殖，雌体把蛋产在底层，没有父母的照看。
由于其他鱼类对季风气候的适应，其繁殖很可能受雨季的到来影响，虽然在旱季观察到成熟雌性的卵提示斯彭斯等人猜测繁殖很可能依赖于可利用的食物资源，这个水平与降雨量成正相关。
鸡蛋是底栖的，一般4-7天孵出幼体。
雄性斑马鱼包围在产卵地点的周围，并且也采取一种选择追逐雌性的交配方法。
雌性的择偶可能与嗅觉有关。
实验室的实验结果表明，斑马鱼幼鱼偏好于利用嗅觉和视觉与亲鱼联系。
这一观察表明，野外成群的幼鱼很可能是近亲，尽管这个还没有被证实。
浅滩成鱼个体的亲缘程度也不知道，尽管实验数据显示个体的喜好与性成熟后避免性逆转有关系，表明在某一点上仔稚鱼远离产后的浅滩。
这个群体遗传学研究支持的前提表明，从印度四个不同的地方收集来的鱼类高水平的遗传变异和薄弱的遗传结构，至少以微卫星标记为依据。
2.3 寿命到目前为止，还没有研究报告详细介绍野外斑马鱼的寿命。
斯彭斯等人推测斑马鱼的寿命主要是一年，以这一年中每月采集样品的标本大小分布为依据，和对其繁殖方式的评价。
然而这些数据是有局限性的，并且只有一个单一的群体。
记录的实验室斑马鱼的寿命可以超过五年。
进一步收集的野生种群的年龄组成将有利于野生的和实验室动物年龄的比较，这将对生物学研究有用处，尤其是那些老龄的。
2.4 食性野生斑马鱼对食物喜好的信息与畜牧业有关，因为它可能有助于饮食和饲养计划的设计，比目前的人工饲养更好的反映特定物种的消化生理和摄食行为。
有一些可利用的数据提供了自然界中斑马鱼的摄食习惯。
斑马鱼是杂食性动物，摄食多种底栖动物和浮游甲壳类动物，以及蠕虫和昆虫幼虫。
他们对双翅目幼虫的摄食倾向被建议用来对蚊虫的控制。
麦克卢尔等人分析了来自印度三个不同采样地点斑马鱼的肠道内容物，发现其摄取的主要是陆地上的一些昆虫。
这些作者认为这种模式是该物种小生境偏好的结果，因为经常发现斑马鱼的池塘和河流也是那些昆虫最容易掉落水中的地方。
虽然这些数据确实表明，在高降雨期间更多的调查研究对获取对自然界中该物种摄食习惯更全面的了解是必要的。
3.斑马鱼文化3.1 水化学斑马鱼作为实验动物模型的产生最主要的因素是它对在范围宽广野外环境生存的耐受力。
它们的适应能力是对它们在野外分布的一个反映，由于当地的地理因素和明显的季节性降雨波动模式导致它们在一系列物理化学性质有很大不同的栖息地被发现的。
然而，重要的是要认识到，养殖最佳环境范围以外的鱼需要比较大的耗费。
最理想条件下养殖动物，必须投入维持平衡越来越多的能量比例，而不是生长，产卵和免疫。
次优条件的结果是生长率，子代数量和质量和最终，生存的下降。
因此，至关重要的是确定人工饲养条件下斑马鱼水质参数的范围，这样可以降低死亡率，使鱼体生长迅速，在性成熟后产下大量高质量的胚胎。
3.1.1 温度温度是鱼类养殖作业时的一个最重要的物理参数之一，因为它在生命历程中的生物化学过程发挥着重要的影响。
....动物，像鱼类，显示其对温度变化不同程度的耐受性，以及它们生存最好的更窄的最佳范围。
斑马鱼可以分为广温性，因为它们表现出更宽的温度耐受性。
从实验室控制的数据表明，斑马鱼对温度的最大耐受范围是6.7-41.7度，这是它与已知的最广温性鱼类之一的希普斯桃花鱼相似。
值得重要注意的是这两项研究中的耐受范围强烈的受到驯化温度的影响；比如说鱼类，在较低温度下驯化一定时间可以延长到更低的温度范围比在较高温度下驯化延长到更高的温度的范围大。
这种温度变化的灵活性在自然种群中也有记录，在孟加拉国九个不同采集地点水温中斑马鱼的观察收集可知其温度范围是16.5-33度。
这些数据提供强有力的证据表明，实验室证明的斑马鱼宽广的耐受性不是人为因素。
斑马鱼的最佳耐热还没有正式确定。
韦斯特在斑马鱼培育中普遍建议维持温度是28.5度，这个最佳温度至少有一项发表的实验研究支持，当斑马鱼在28度时表现出最快的增长速度，而不是较低和∕或随机的和人为的温度波动。
然而，以其在自然界中的广泛耐受性为基础，它实际的喜好温度高于或低于这个温度范围是极有可能的，马修斯等人建议的24-30度的温度范围可能更合适。
尽管对韦斯特建议的一般坚持已经导致温度是斑马鱼研究和养殖中最重要的环境参数，应该认识到在喜好温度之外的控制和饲养温度的偏差可能在许多方面的研究中造成影响。
饲养温度的升高已经被证明扰乱了调节性分化的类固醇合成酶的合成，从而使实验室种群性别比倾斜。
通过分子蛋白变温来维持细胞的功能效率比较低，从而在最适温度以外影响一些鱼类的表现。