鸡PPAR基因单核苷酸多态与脂肪性状相关的研究

以AA肉鸡和 3个中国地方鸡品种 (石岐杂、北京油鸡、白耳鸡 )为实验材料 ,用 7对引物对PPAR α基因整个编码区用PCR SSCP方法进行了扫描 ,结果发现引物 4扩增片段上有一个C T的单碱基突变 ,使鸡群中表现出 3种基因型 (AA、AB、BB)。统计结果发现 3种基因型在各品种中的分布不一致 ,AA肉鸡同中国地方鸡品种差异很大 ,χ2 独立性检验差异极显著 (P <0 0 1)。将AA肉鸡 3种基因型同腹脂等屠体性状进行方差分析 ,结果表明BB基因型与其他 2种基因型相比有较高的腹脂重和腹脂率 ,同AA型比较差异显著 (P <0 0 5 )。因此推测PPAR α基因可能是影响鸡脂肪代谢的主效基因或与控制该性状的主效基因连锁 ,并且能够用于对鸡脂肪性状进行分子标记辅助选择。     

鸡Myostatin基因单核苷酸多态性的群体遗传学分析

肌肉生长抑制素是控制骨骼肌生长发育的重要细胞因子,采用PCR－SSCP和测序的方法发现了5个位于Myostatin基因5′－和3′－调控区的单核苷酸多态性位点,对北京油鸡、白耳鸡、石歧杂、矮小黄鸡、小型黄鸡、惠阳胡须鸡、隐性白羽鸡、海兰、AA鸡等不同鸡种的该单核苷酸多态性分析结果表明：Myostatin基因的5′调控区引物P60／P61扩增片段多态性是由3个核苷酸的改变而产生的[分别是G→A（304位）、A→G（322位）、G→（344位）],引物P93／P94扩增片段的多态性是由G→A（167位）突变造成的,引物P117。P118PC扩增片段多态性是由T→C（177位）造成的。3′调控我引物P80／P81扩增片段多态性是由第7263位A突变为T造成的,引物P76／P77扩增片段多态性是由A→G（6935位）造成的。不同鸡种群体遗传学分析表明,5′－调控区引物60／P61扩增片段多态性片段多态性是由A→G（6935位）造成的。不同鸡种群体遗传学分析表明,5′－调控区引物P60／P61扩增片段多态性位点在北京油鸡的基因型频率分布与其他的品种有很大的差异,其BB型频率为0．700,AA基因型频率仅为0．033,而其他鸡种中以A基因优势;对于引物P93／P94,品种间的基因型频率差异极显著（P＜0．01）,北京油鸡和AA鸡的EE型频率鸡种中以A基因占优势;对于引物P93／P94,品种间的基因型频率差异极显著（P＜0．01）,北京油鸡和AA鸡的EE型频率低于其他品种,白耳鸡和海兰蛋鸡以EE型为主,其频率高于其他品种;3′－调控区引物P80／P81多态怀位点在9个鸡种中都是等位基因C占优势。引物P76／P77,总体上MM型的频率较低,杂合子MN型的频率较高。     

鸡瘦蛋白受体(OBR)基因外显子9单核苷酸多态性分析

瘦蛋白受体属于Ⅰ类细胞因子超家族受体,其在瘦蛋白的信号转导中起着关键的作用。本研究根据瘦蛋白受体基因外显子9的序列设计引物,用PCR-SSCP的方法对高脂系（fatness line,FL）、低脂系(leanness line,LL)、北京油鸡、白耳鸡、石歧杂、矮小黄鸡、小型黄鸡、惠阳胡须鸡、隐性白羽鸡和海兰蛋鸡等品种（系）进行了单核苷酸多态性分析,并检测到了多态性。对两种纯合子片段克隆和测序,结果表明在编码区的1167位碱基发生了突变,由C突变为A,但是编码的氨基酸并没有发生改变。经独立性检验,基因型频率和基因频率分布与品种有关,北京油鸡的AA基因型频率显著高于其他品种;高脂系中A基因频率显著高于低脂系。 Abstract:Leptin receptor is a type I cytokine super family member and plays an important role in leptin functioning signal transduction.This study was designed to investigate the single nucleotide polymorphism (SNP) of OBR gene in various breeds,including Fatness Line (FL),Leanness Line (LL),Beijing Youji,Baierji,Shiqiza,Dwarf Yellow Chickens,Mini Yellow Chickens,Huiyang Huxuji,Recessive White Chickens and Hyline Layer.The primers for exon 9 in OBR gene were designed from the database of chicken genomic sequence and the SNPs were detected by PCR-SSCP method.One SNP (C/A at 1167 in cds) was found among individuals within all breeds.However,the amino acid was not changed because it was a silence mutantion.The result of population genetics analyses showed that the frequency of AA genotype in Beijing Youji was significantly higher than that in other lines.Also,the frequency of A allele in FL was significantly higher than that in LL.     

鸡A-FABP基因多态性分析及其与脂肪性状的

以北京油鸡为试验材料,对A-FABP基因进行单核苷酸多态性(SNPs)检测和基因型与性状的关联分析。方差分析结果表明,不同基因型间腹脂率、皮脂厚、肌内脂肪含量差异极显著(P<0.01),体重在不同基因型间差异不显著(P >0.05)。由此推测,A-FABP可能为影响鸡脂肪代谢的主效基因或与主效基因相连锁。     

鸡瘦蛋白受体(OBR)基因内含子8单核苷酸多态性与体脂性状的相关研究

以东北农业大学高低脂双向选择系的第 6世代肉鸡为材料 ,鸡 7周龄时测定体重和腹脂重等屠体性状。根据鸡瘦蛋白受体基因内含子 8的序列 (GenBank登陆号 :AF2 2 2 783 )设计引物 ,用直接测序的方法检测多态性位点 ,用PCR SSCP的方法进行基因型分析 ,建立适合的统计模型对多态性位点产生的基因型与生长和体组成性状进行相关分析。结果表明 ,在第 50 0和 659位碱基同时发生了T—C、G—A突变。经最小二乘分析 ,3种基因型在腹脂重和腹脂率上差异显著 (P <0 0 5) ,BB型个体腹脂重和腹脂率显著高于AB型 (P <0 0 5) ,极显著地高于AA型个体 (P <0 0 1) ;3种基因型在肝重上差异显著 (P <0 0 5) ,且AA基因型个体的肝重显著低于AB和BB基因型个体。初步推断OBR基因可能是影响鸡脂肪性状的主效基因或与主效基因连锁 ,推测可以利用这个多态位点对鸡的体脂性状进行标记辅助选择     

钙蛋白酶Ⅰ(CAPN1)基因多态性与鸡肉嫩度和屠体性状的相关研究

为了探讨CAPN1基因作为影响鸡肌肉嫩度候选基因的可能性, 寻找与鸡嫩度性状相关的分子标记, 对钙蛋白酶Ⅰ(CAPN1)基因的CDS区进行SNPs 检测, 分析不同基因型在5个优质肉鸡纯品系和3个配套系间分布规律。利用测序和单链构象多态(SSCP)的方法进行SNPs 检测和基因型的分析, 计算等位基因频率、各位点多态信息含量。结果发现2546位(位点A) 处发生点突变由C→T和3535位(位点B)处发生点突变由G→A。各位点的3 种基因型与肉鸡生产性状的最小二乘分析结果表明,各位点的各种基因型个体在肌纤维密度和部分屠体性状指标存在显著差异(P< 0.05)。初步推断CAPN1基因可能是影响鸡嫩度性状潜在的主效基因或与主效基因连锁, 并且这些位点具有成为分子标记的潜在可能。     

脂肪分化相关蛋白基因多态性与优质鸡屠宰性状和肌内脂肪含量的相关性研究

对脂肪分化相关蛋白(Adipocyte Differentiation-Related Protein, ADFP)基因的外显子进行SNPs 检测, 探讨其作为鸡脂肪性状候选基因的可能性。实验以四川省畜牧科学研究院和大恒家禽育种有限公司培育的优质肉鸡新品系为素材, 采用PCR-SSCP的方法进行SNPs 检测和基因型的分析。结果找到3个SNPs位点: 4 079位由A→T(位点A)、4 843位由C→T(位点B)和7 070位由A→G(位点C)。单位点基因型对屠宰性状的遗传效应分析表明, 位点A的基因型对腿肌率、腹脂重、腹脂率和肌内脂肪含量有显著性影响(P < 0.05), 位点B的基因型对活重和屠体重均有显著性影响(P < 0.05), 位点C的基因型对胸肌重和肌内脂肪含量有显著性影响(P < 0.05), 对胸肌率有极显著性影响(P < 0.01)。初步推断ADFP基因可能是影响鸡脂肪性状的主效基因或与主效基因连锁, 推测可以利用多态位点A和C对鸡腹脂重、腹脂率和肌内脂肪含量进行标记辅助选择。     

利用基因芯片技术研究两品种鸡脂肪组织差异表达基因

应用包含9024条鸡cDNA的表达谱芯片,对从两品种鸡脂肪组织抽提及纯化的mRNA进行芯片杂交,并对基因表达谱进行分析,旨在筛选高脂肉鸡和白耳蛋鸡脂肪组织差异表达的基因,探讨造成两品种体脂性状差异的分子生物学机理。结果按差异显著阳性标准分析,共筛选出差异表达基因67条,主要涉及脂类代谢、能量代谢、细胞骨架构成、转录和剪接因子以及蛋白质合成与分解等相关基因,此外,还筛选出一些尚未在GenBank上登陆的序列,推测可能是未知的新基因,它们在鸡脂类代谢的过程所起到的作用还需进一步实验证明。     

鸡生长激素基因5‘端部分调控区的克隆分析

利用ＰＣＲ技术扩增、克隆、测序了７个跨越不同生长速度的鸡品种（系）的生长激素（ＧＨ）基因的５＇端部分调控区。这７个品种（系）分别为：高生长速度的宝罗肉鸡父本;较高生长速度和一定的产蛋性能的宝罗肉鸡母本;肉蛋灵用型的芦花鸡;中等生长速度和中等体重的蛋鸡品种洛岛红和农大褐;生长速度较慢体重较轻的蛋鸡品种北京白鸡和生长速度很慢但又不是矮小型的地方品种丝毛乌骨鸡。所扩增的片段长度为７６０ｂｐ,包括了转录起     

A-FABP基因多态性与肉鸡生长和体组成性状的关联

为探讨A-FABP基因多态性对肉鸡生长和体组成性状的影响,文章选用肉鸡高、低脂双向选择品系第十世代肉仔鸡为实验材料,采用测序、PCR-RFLP、PCR-LP、DHPLC方法进行基因多态性检测和个体基因型分析,通过对8个SNPs进行连锁不平衡分析并选择5个标签SNPs构建单倍型,进而利用单位点和单倍型分别与鸡生长和体组成性状进行关联分析。结果表明,7个SNPs(除SNP 5以外)及单倍型同时对鸡肌胃重、肌胃率有显著影响(P<0.05),而对生长和其他体组成性状无显著影响(P>0.05)。鉴于不同物种A-FABP基因现有的功能研究均没有发现其对消化系统有重要作用,因此该基因是否为影响肉鸡肌胃重和肌胃率的主效基因还有待于进一步研究。     

Gene Duplication and Gene Conversion in the Caenorhabditis elegans Genome

A comprehensive analysis of duplication and gene conversion for 7394 Caenorhabditis elegans genes (about half the expected total for the genome) is presented. Of the genes examined, 40% are involved in duplicated gene pairs. Intrachromosomal or cis gene duplications occur approximately two times more often than expected. In general the closer the members of duplicated gene pairs are, the more likely it is that gene orientation is conserved. Gene conversion events are detectable between only 2% of the duplicated pairs. Even given the excesses of cis duplications, there is an excess of gene conversion events between cis duplicated pairs on every chromosome except the X chromosome. The relative rates of cis and trans gene conversion and the negative correlation between conversion frequency and DNA sequence divergence for unconverted regions of converted pairs are consistent with previous experimental studies in yeast. Three recent, regional duplications, each spanning three genes are described. All three have already undergone substantial deletions spanning hundreds of base pairs. The relative rates of duplication and deletion may contribute to the compactness of the C. elegans genome. Received: 30 July 1998 / Accepted: 12 October 1998     

MADS-box基因家族基因重复及其功能的多样性

基因的重复(duplication)及其功能的多样性(diversification)为生物体新的形态进化提供了原材料。MADS-box基因在植物(特别是被子植物)的进化过程中发生了大规模的基因重复事件而形成一个多基因家族。MADS-box基因家族的不同成员在植物生长发育过程中起着非常重要的作用,在调控开花时间、决定花分生组织和花器官特征以及调控根、叶、胚珠及果实的发育中起着广泛的作用。探讨MADS-box基因家族的进化历史有助于深入了解基因重复及随后其功能分化的过程和机制。本文综述了MADS-box基因家族基因重复及其功能分化式样的研究进展。     

Gene Function Prediction Based on the Gene Ontology Hierarchical Structure

The information of the Gene Ontology annotation is helpful in the explanation of life science phenomena, and can provide great support for the research of the biomedical field. The use of the Gene Ontology is gradually affecting the way people store and understand bioinformatic data. To facilitate the prediction of gene functions with the aid of text mining methods and existing resources, we transform it into a multi-label top-down classification problem and develop a method that uses the hierarchical relationships in the Gene Ontology structure to relieve the quantitative imbalance of positive and negative training samples. Meanwhile the method enhances the discriminating ability of classifiers by retaining and highlighting the key training samples. Additionally, the top-down classifier based on a tree structure takes the relationship of target classes into consideration and thus solves the incompatibility between the classification results and the Gene Ontology structure. Our experiment on the Gene Ontology annotation corpus achieves an F-value performance of 50.7% (precision: 52.7% recall: 48.9%). The experimental results demonstrate that when the size of training set is small, it can be expanded via topological propagation of associated documents between the parent and child nodes in the tree structure. The top-down classification model applies to the set of texts in an ontology structure or with a hierarchical relationship.     

Gene Circuit Analysis of the Terminal Gap Gene huckebein

The early embryo of Drosophila melanogaster provides a powerful model system to study the role of genes in pattern formation. The gap gene network constitutes the first zygotic regulatory tier in the hierarchy of the segmentation genes involved in specifying the position of body segments. Here, we use an integrative, systems-level approach to investigate the regulatory effect of the terminal gap gene huckebein (hkb) on gap gene expression. We present quantitative expression data for the Hkb protein, which enable us to include hkb in gap gene circuit models. Gap gene circuits are mathematical models of gene networks used as computational tools to extract regulatory information from spatial expression data. This is achieved by fitting the model to gap gene expression patterns, in order to obtain estimates for regulatory parameters which predict a specific network topology. We show how considering variability in the data combined with analysis of parameter determinability significantly improves the biological relevance and consistency of the approach. Our models are in agreement with earlier results, which they extend in two important respects: First, we show that Hkb is involved in the regulation of the posterior hunchback (hb) domain, but does not have any other essential function. Specifically, Hkb is required for the anterior shift in the posterior border of this domain, which is now reproduced correctly in our models. Second, gap gene circuits presented here are able to reproduce mutants of terminal gap genes, while previously published models were unable to reproduce any null mutants correctly. As a consequence, our models now capture the expression dynamics of all posterior gap genes and some variational properties of the system correctly. This is an important step towards a better, quantitative understanding of the developmental and evolutionary dynamics of the gap gene network.     

腺病毒载体介导CEA基因元件调控自杀基因治疗

构建以CEA启动子控制HSV-TK基因表达的复制缺陷型腺病毒载体(AdCEATK).纯化的重组腺病毒滴度达1×10¹²pfu/ml.CEA阴性的HeLa细胞感染AdCMVTK后对丙氧鸟苷(GCV)很敏感,而感染了AdCEATK后不被GCV杀伤.与此相反CEA阳性的LoVo细胞中AdCMVTK和AdCEATK都有很好的表达活性,说明CEA启动子有良好的细胞专一性.AdCEATK/GCV系统还有明显的旁杀伤效应.此载体将有助于实现对CEA阳性肿瘤的专一性自杀基因治疗.     

Transient Gene Expression in Tobacco using Gibson Assembly and the Gene Gun

In order to target a single protein to multiple subcellular organelles, plants typically duplicate the relevant genes, and express each gene separately using complex regulatory strategies including differential promoters and/or signal sequences. Metabolic engineers and synthetic biologists interested in targeting enzymes to a particular organelle are faced with a challenge: For a protein that is to be localized to more than one organelle, the engineer must clone the same gene multiple times. This work presents a solution to this strategy: harnessing alternative splicing of mRNA. This technology takes advantage of established chloroplast and peroxisome targeting sequences and combines them into a single mRNA that is alternatively spliced. Some splice variants are sent to the chloroplast, some to the peroxisome, and some to the cytosol. Here the system is designed for multiple-organelle targeting with alternative splicing. In this work, GFP was expected to be expressed in the chloroplast, cytosol, and peroxisome by a series of rationally designed 5’ mRNA tags. These tags have the potential to reduce the amount of cloning required when heterologous genes need to be expressed in multiple subcellular organelles. The constructs were designed in previous work¹¹, and were cloned using Gibson assembly, a ligation independent cloning method that does not require restriction enzymes. The resultant plasmids were introduced into Nicotiana benthamiana epidermal leaf cells with a modified Gene Gun protocol. Finally, transformed leaves were observed with confocal microscopy.     

Gene targeting in the mouse

Mice with alterations to specific endogenous genes can be produced by gene targeting in embryonic stem cells. The field has developed rapidly over the past decade, so that large numbers of mice with different gene deficiencies have been generated. Knockout mice provide an ideal opportunity to analyse the function of individual mammalian genes and to model a range of human inherited disorders. This powerful approach has also identified numerous examples of gene redundancy and has highlighted the need to consider metabolic differences between man and mouse in disease modelling. More sophisticated gene-targeting methods are now being used to introduce subtle gene alterations. In the future, more refined genetic analysis and genome, rather than individual gene, alterations will be achieved by incorporating site-specific recombination into targeting strategies. Gene targeting could also make a contribution to improved protocols for gene therapy.