Characterization of expressed sequence tags from a gallus gallus pineal gland cDNA library

The pineal gland is the circadian oscillator in the chicken, regulating diverse functions ranging from egg laying to feeding. Here, we describe the isolation and characterization of expressed sequence tags (ESTs) isolated from a chicken pineal gland cDNA library. A total of 192 unique sequences were analysed and submitted to GenBank; 6% of the ESTs matched neither GenBank cDNA sequences nor the newly assembled chicken genomic DNA sequence, three ESTs aligned with sequences designated to be on the Z_random, while one matched a W chromosome sequence and could be useful in cataloguing functionally important genes on this sex chromosome. Additionally, single nucleotide polymorphisms (SNPs) were identified and validated in 10 ESTs that showed 98% or higher sequence similarity to known chicken genes. Here, we have described resources that may be useful in comparative and functional genomic analysis of genes expressed in an important organ, the pineal gland, in a model and agriculturally important organism.     

中国红原鸡和泰国红原鸡遗传多样性分析

利用29个微卫星DNA标记对来自中国的红原鸡Gallus gallus spadiceus亚种和来自泰国的红原鸡Gallus gallus gallus亚种进行遗传多样性分析, 评估亚种内的遗传变异和亚种间的遗传分化, 结果表明: 共检测到168个等位基因, 每个位点的等位基因数从2到13不等, 所有位点平均的期望杂合度和PIC值分别为0.5780和0.53。中国和泰国红原鸡29个微卫星位点平均有效等位基因数分别为3.79和4.79, 平均基因杂合度为0.5379和0.6385, 两个红原鸡亚种均表现出较高的群体杂合度和丰富的遗传多样性。群体分化系数为19.4%(P<0.01), 两个红原鸡亚种间的Reynolds’遗传距离和Nm值分别为0.157和1.040。由此可见, Gallus gallus spadiceus亚种和Gallus gallus gallus亚种群体具有不同的群体遗传结构, 群体之间存在明显的遗传分化, 并不能将其认定为是同一亚种, 这也为中国家鸡具有独立的起源提供了一定的佐证。     

Genetic characterization of Bhutanese native chickens based on an analysis of Red Junglefowl (Gallus gallus gallus and Gallus gallus spadecieus), domestic Southeast Asian and commercial chicken lines (Gallus gallus domesticus)

The genetic diversity of Bhutanese chickens needs to be understood in order to develop a suitable conservation strategy for these birds in Bhutan. In this, work, we used microsatellite markers to examine the genetic diversity of Bhutanese chickens. Four Bhutanese chicken varieties (Black plumage, Frizzle, Naked neck and Red Junglefowl-like, corresponding to Yuebjha Narp, Phulom, Khuilay and Seim, respectively), two subspecies of Red Junglefowl (Gallus gallus gallus and Gallus gallus spadecieus), two varieties of Thai native chickens (Pradhu Hang Dam and Chee; Gallus gallus domesticus) representing the Southeast Asian domestic chicken, and two commercial lines (Broiler and Single Comb White Leghorn) were genotyped with 18 microsatellites that included 16 loci recommended by the FAO/ISAG for investigations of genetic variability in chickens. All loci were polymorphic, with the number of alleles ranging from six (MCW0111) to 23 (MCW0183). Substantial genetic variation was observed in all populations, with the Bhutanese native chicken Yuebjha Narp (Black plumage chicken) showing the lowest genetic variability. Despite extensive intrapopulation variation, the genetic differentiation among 10 populations was moderate. A neighbor-joining tree revealed the genetic relationships involved while principal component analysis showed that Bhutanese native chickens should be given priority in conservation efforts because of their genetic distinctiveness. Chee chickens are especially valuable as a reservoir of predomestic diversity, as indicated by their greater genetic variation and their position in the phylogenetic tree.     

Crystallization and preliminary x-ray analysis of ovocleidin-17 a major protein of the gallus gallus eggshell calcified layer

In this work, we report the crystallization of ovocleidin-17, the major protein of the avian eggshell calcified layer and the preliminary X-ray characterization of this soluble protein which is implied into the CaCO(3) formation of the eggshell in avians. Crystals belong to one of the trigonal space group P3 with cell dimensions a= b= 59.53 A and c = 83.33 A, and alpha=beta= 90 degrees and gamma=120 degrees. Crystals diffract up to 3.0 A.     

Investigation of the antigen recognition and presentation capacity of pecteneal hyalocytes in the chicken (gallus gallus domesticus)

We investigated at different developmental stages the antigen recognition and presentation capacity of pecteneal hyalocytes that adhere to the pecten oculi in the eye of domestic chickens. Forty-eight fertilized eggs were used to investigate embryonic stages and 12 6-week-old chickens were used to investigate adults. Tissue samples from both embryos and adults were stained with monoclonal antibodies against MHC-II, TLR2/CD282 and TLR4, and also with RCA-1, WGA and SNA lectins. The developmental stage of the pecteneal hyalocytes was determined using Masson's triple staining. Pecteneal hyalocytes first appeared at Hamburger-Hamilton stages 30 34 and remained unchanged from their first appearance to adulthood. Chicken pecteneal hyalocytes were stained by monoclonal antibodies against TLR2 and TLR4, and were unstained by monoclonal antibodies for MHC-II. Hyalocytes were positive for RCA-I, WGA and SNA lectins. We found that pecteneal hyalocytes that adhere to the pecten oculi in domestic chickens recognized antigens, but could not present them. These cells have been assumed to be of monocyte/macrophage lineage; their functions in the immune response are not fully understood.     

Ultrastructural characteristics of anterior gut innervation of Gallus gallus

The enteric nervous system of the bird's anterior gut is very well developed. Myelin fibres are seen accompanying the nervous trunks up to the mucous layer. Glial cells duplicate the number of neurons in the myenteric plexuses. Their number decreases at the submucous plexuses, but it is always higher than the neurons. Isolated neurons are widely spread in the circular muscle coat accompanying the nervous trunks which can be inter and intrafascicularly located. Direct synaptic contacts with the soma neuronal membranes are very often seen. We have never observed synaptic specializations. The most prominent varicosities either in the peripheric nervous trunk axons or directly laying on the soma membranes are those containing peptidergic or mixed vesicles of cholinergic and peptidergic types. The neurons show big nuclei of different size and shape. Neighbouring smooth muscle cells show abundant caveolae near the nervous elements. Although we have not observed close contacts with glands, thin axon bundles spread near the glandular cells of the mucous layer.     

Parameters of the chicken genome (Gallus gallus)

As more information on the chicken genome is gathered, it is becoming increasingly more important to be able to correlate genetic and physical maps. Quantitation of the chicken karyotype is important in establishing parameters which define the genome. Here we report on the physical lengths of the chicken macrochromosomes and establish the DNA content of each, thus identifying implicitly how much of the genome is represented by the microchromosomal component. For the first time, genetic and physical data on the chicken karyotype are presented in relation to one another.     

红原鸡全基因组中微卫星分布规律研究

本文对红原鸡Gallus gallus全基因组中微卫星数量及分布规律进行了分析,查找到l～6个碱基重复类型的微卫星序列共282728个,约占全基因组序列(1.1Gb)的0.49%,分布频率为1/3.89kb,微卫星序列的长度主要在12～70个碱基长度范围内。第1、2、3条染色体上微卫星分布频率较高,而32号染色体上无微卫星分布。不同类型微卫星中,单碱基重复类型数目最多,为184192个,占总数的65.1%;其次是四、二、三、五、六碱基重复单元序列,分别占到总数的12.8%、9.7%、7.2%、4.6%、0.8%。T、A、AT、GTTT、AAAC、G、C、ATTT、AC、GT、AAAT、ATT、AAC、AAT、GTT、AG、CT、CTTT、AAAG、GTTTT、AAACA、AAGG、CCTT是红原鸡基因组中最主要的微卫星重复类型。本研究为红原鸡微卫星标记的分离筛选、遗传多样性的研究以及不同物种微卫星的比较分析奠定了基础。     

Functional morphology of the glomerular filtration barrier of Gallus gallus

The anionic charge barrier and the endothelial and epithelial pore sizes on the glomerular filtration barrier (GFB) were examined in white leghorn chickens (Gallus gallus). Ruthenium red was used to stain anionic charge sites on the GFB. The tissue was treated by normal dehydration and freeze substitution dehydration for transmission electron microscopy (TEM). In addition, the basal lamina was isolated for study. The results of our study indicate that G. gallus possess a thick, negatively charged glycocalyx surrounding the podocytes and slit diaphragm and on the endothelium. However, in all cases, little anionic charge is present in the basal lamina. The pores on the endothelium are elliptical and have mean dimensions of 148 × 110 nm. This is in contrast to mammals, which have smaller, round pores. The epithelial pores in G. gallus measure approximately 35 nm in length, approximately 4 times larger than those found in mammals. These results indicate that the avian glomerulus may allow the filtration of larger molecules from the plasma than occurs in mammals and that the charge on the molecule may not be as restrictive a filtration characteristic as in mammals. © 1996 Wiley-Liss, Inc.     

原鸡海南亚种的巢址特征

选择适宜的巢址对降低巢捕食风险,提高繁殖成效有重要意义。2008年3—7月,在海南省大田国家级自然保护区,采用随机样线法结合访问法在各种生境类型中系统地寻找原鸡海南亚种(Gallus gallus jabouillei)的巢,并在野外追踪观察基础上用样方法对原鸡的巢址选择进行研究。共记录到原鸡的巢16个,以巢为中心选取样方并测量反映巢址的13个特征生境参数,同时分析16个对照样方以进行比较,并通过主成分分析探讨影响巢址选择的主导因子。结果表明:(1)原鸡的巢位于密集的丛生草本下或低矮的灌木丛基部。营巢背景多样,其中位于草本植物下10个(62.5%);灌草丛5个(31.3%);灌木下1个(6.25%)。(2)Mann–WhitneyU检验的结果显示,原鸡偏好于比较开阔、地面落叶稀少而靠近的小道和林缘的位置营巢。(3)主成分分析表明,植被盖度因子和空间位置因子贡献率最大,在降低巢捕食风险中有重要价值,是影响原鸡巢址选择的主要因素。     

The primary sequence of chicken myoglobin (Gallus gallus).

After enzymatic digestion of chicken myoglobin by trypsin, chymotrypsin or thermolysin, the separation of peptides was performed by column chromatography on various ion exchange resins. Each peptide was purified by high-voltage paper electrophoresis or by chromatography either on paper or on ion-exchange resin, and its complete amino acid sequence was then determined by the combined dansyl-Edman procedure and by endopeptidase digestions. The whole globin was submitted to automatic Edman degradation using the Beckman sequencer. Residues have been positioned from overlaps of sequence data between tryptic (T), chymotryptic (C) and thermolysin (Th) peptides. The stepwise degradation of the whole globin confirmed the alignment of the N-terminal third of the molecule. The combination of these different approaches has led to the complete determination of the 153 residues sequence forming the polypeptide chain of chicken myoglobin. Comparison of the established chicken myoglobin structure with those from other species shows a conservation of structure, although the avian protein exhibits more variations in its amino acid sequence than has been found between other known myoglobins which all belong to mammalian species.     

Material on experimental polyembryony in Gallus gallus domestica chickens

Cutting the blastodisc of nonincubated chick eggs in parallel and perpendicularly to the presumptive axis results in the development of twin embryos in 28% of cases. After tripsin and versen solutions were injected in various concentrations into the blastula cavity, the part of the developed twins was but negligible (5% and 3%, respectively). In all experimental groups embryos with different developmental anomalies were often occurrence, as well as various types of malformation were noted in twins. Histological investigation of embryos having teratomic character demonstrated double anlage of some organs in them. Formation of several neural tubes, chordae and hearts was noted. It is possible that in most cases somatic embryogenesis takes place, incompleted due to intensive processes of regulatory character at early embryogenesis in avian. Results of in vitro experiments for studying regulatory capability of early chicken embryos (mechanical trauma was performed in the embryo for this purpose) demonstrate a rather high degree of integration that should prevent processes of polyembryony to develop in the experiments.     

原鸡海南亚种的巢址特征

选择适宜的巢址对降低巢捕食风险,提高繁殖成效有重要意义。2008年3—7月,在海南省大田国家级自然保护区,采用随机样线法结合访问法在各种生境类型中系统地寻找原鸡海南亚种(Gallus gallus jabouillei)的巢,并在野外追踪观察基础上用样方法对原鸡的巢址选择进行研究。共记录到原鸡的巢16个,以巢为中心选取样方并测量反映巢址的13个特征生境参数,同时分析16个对照样方以进行比较,并通过主成分分析探讨影响巢址选择的主导因子。结果表明：（1) 原鸡的巢位于密集的丛生草本下或低矮的灌木丛基部。营巢背景多样,其中位于草本植物下10个(62.5%);灌草丛5个(31.3%);灌木下1个(6.25%)。（2) Mann–Whitney U检验的结果显示,原鸡偏好于比较开阔、地面落叶稀少而靠近的小道和林缘的位置营巢。（3) 主成分分析表明,植被盖度因子和空间位置因子贡献率最大,在降低巢捕食风险中有重要价值,是影响原鸡巢址选择的主要因素。     

EST analysis of mRNAs expressed during embryogenesis in Gallus gallus

Chicken Expressed Sequence Tags (ESTs) were analyzed to identify genes associated with myogenesis during embryonic development. A total of 6,184 ESTs were generated from three cDNA libraries constructed from whole embryos (Stage 26), somites associated with neural tube (Stage 15), and limb buds (Stages 21, 24 and 26). Clustering and assembly of 4,998 valid ESTs resulted in 2,329 unique sequences with 902 clusters (38.7%) and 1,427 singletons (61.3%). There are more than 400,000 chicken ESTs available at GenBank and we were able to identify 143 novel sequences. From these, 45 sequences found either a human EST homolog or a match with conserved regions among proteins. Most of these sequences were found to be expressed in somites, an important tissue for muscle development and not characterized before. This study revealed the value of micro dissected embryonic libraries for describing gene expression profiles associated with myogenesis and gene discovery.     

Molecular diagnosis of Eimeria species affecting naturally infected Gallus gallus

We used PCR to test various protocols and define a technique for DNA extraction directly from chicken-shed stool samples for the identification of Eimeria species that parasitize birds. It was possible to extract and amplify DNA of seven Eimeria species from field stool samples, using both protocols tested; extractions made with phenol/chloroform protocols gave the best results. The primers were specific and sensitive, allowing amplification of samples containing as few as 20 oocysts, both in individual and in a multiplex PCR. Individualized PCR with the phenol/chloroform DNA extraction protocol detected a larger number of Eimeria species. Molecular diagnosis was found to be practical and precise, and can be used for monitoring and epidemiological studies of Eimeria.