Search

In order to breed a strain that has heat tolerance and meat productivity, the commercial red-feathered Taiwan native chickens were male (F group), and heat stress resistant strain Taiwan native chickens (Taishu-9, bred by the Taiwan Livestock Research Institute) were female (TR9 group) to hybridize to generate offspring (F9 group). Three breeds of birds (male) were conducted to compare acute heat stress and meat quality. At 12 weeks of age, TR9 group showed the significantly lowest activity of plasma creatine kinase upon acute heat stress which indicated heat stress resistant in TR9 group as expected. In addition, only limited thermoregulation was obtained in F9 group, while F group exhibited almost no acute heat stress tolerance ability. After slaughtered at 16 weeks of age, the F group revealed poor meat quality in breast meat as pale, soft, and exudative (PSE)-like muscle samples according to CIE L* and pH value. The F9 group was an offspring of TR9 group with heat tolerance, but it only demonstrated limitation of heat resistance. However, the improve meat quality was obtained in F9 group compared to F group, and that may be contributed from better anti-stress as like as TR9 group during slaughtering process.

Background Metastasis is the major cause of morbidity and mortality in cancer that involves in multiple steps including epithelial–mesenchymal transition (EMT) process. Centrosome is an organelle that functions as the major microtubule organizing center (MTOC), and centrosome abnormalities are commonly correlated with tumor aggressiveness. However, the conclusive mechanisms indicating specific centrosomal proteins participated in tumor progression and metastasis remain largely unknown. Methods The expression levels of centriolar/centrosomal genes in various types of cancers were first examined by in silico analysis of the data derived from The Cancer Genome Atlas (TCGA), Gene Expression Omnibus (GEO), and European Bioinformatics Institute (EBI) datasets. The expression of STIL (SCL/TAL1-interrupting locus) protein in clinical specimens was further assessed by Immunohistochemistry (IHC) analysis and the oncogenic roles of STIL in tumorigenesis were analyzed using in vitro and in vivo assays, including cell migration, invasion, xenograft tumor formation, and metastasis assays. The transcriptome differences between low- and high-STIL expression cells were analyzed by RNA-seq to uncover candidate genes involved in oncogenic pathways. The quantitative polymerase chain reaction (qPCR) and reporter assays were performed to confirm the results. The chromatin immunoprecipitation (ChIP)-qPCR assay was applied to demonstrate the binding of transcriptional factors to the promoter. Results The expression of STIL shows the most significant increase in lung and various other types of cancers, and is highly associated with patients’ survival rate. Depletion of STIL inhibits tumor growth and metastasis. Interestingly, excess STIL activates the EMT pathway, and subsequently enhances cancer cell migration and invasion. Importantly, we reveal an unexpected role of STIL in tumor metastasis. A subset of STIL translocate into nucleus and associate with FOXM1 (Forkhead box protein M1) to promote tumor metastasis and stemness via FOXM1-mediated downstream target genes. Furthermore, we demonstrate that hypoxia-inducible factor 1α (HIF1α) directly binds to the STIL promoter and upregulates STIL expression under hypoxic condition. Conclusions Our findings indicate that STIL promotes tumor metastasis through the HIF1α-STIL-FOXM1 axis, and highlight the importance of STIL as a promising therapeutic target for lung cancer treatment.

BACKGROUND Reactive oxygen species (ROS) overproduction is highly related to some human chronic diseases. There are approximately 400 metric tons of chalazae produced yearly after the processing of the liquid-egg production, which are disposed of as waste. The objectives of this study were to look for the optimal production condition of antioxidant crude chalaza hydrolysates and evaluate the in vivo antioxidant capacity via a chronic alcohol consumption mouse model. RESULTS Antioxidant crude chalaza hydrolysates (CCH-As) could be produced by protease A at 1:100 ratio (w/w) and 0.5 h hydrolytic period. After our analyses, CCH-As were rich in leucine, arginine, phenylalanine, valine, lysine and antioxidant dipeptides (anserine and carnosine), and the major molecular masses were lower than 15 kDa. Regarding protective effects of CCH-As against oxidative damage in alcoholic-liquid-diet-fed mice, alcohol-fed mice had lower (P < 0.05) liver antioxidant capacities, and higher (P < 0.05) liver lipid contents, serum lipid/liver damage indices and IL-1β/IL-6 values. CCH-A supplementation reversed (P < 0.05) liver antioxidant capacities and reduced (P < 0.05) serum/liver lipids in alcohol-fed mice, which may result from increased (P < 0.05) fecal lipid output, upregulated (P < 0.05) fatty acid β-oxidation and downregulated (P < 0.05) lipogenesis in the liver. CONCLUSION Taken together, this CCH-A should benefit the liquid-egg industry, while also offering consumers a choice of healthy ingredients from animal sources. © 2018 Society of Chemical Industry.

Four-hundred metric-ton chalazae are produced annually from the liquid-egg processing and always cause a heavy burden due to handling cost in Taiwan. After chalazae were hydrolyzed by protease A, the amounts of hydrophobic, aromatic, and branched-chain amino acids, as well as anserine were dramatically increased. This study was to understand the antifibrogenic effects of protease A-digested crude chalaza hydrolysates (CCH-As) on livers of thioacetamide (TAA) treated rats. CCH-As improved (P< 0.05) growth performance, serum liver damage indices, histopathological liver inflammation, and liver collagen deposition in TAA-treated rats. The antifibrogenic effects of CCH-As were due to decreased (P < 0.05) inflammatory/fibrogenic cytokine contents, α-smooth-muscle-actin (α-SMA) protein expression, and matrix metallopeptidase (MMP)-2 and -9 activities, as well as increased (P < 0.05) the antioxidant capacity in livers. CCH-As also increased (P < 0.05) cleaved caspase-3 and cleaved poly ADP-ribose polymerase protein levels in livers of TAA-treated rats which accelerating cell renewal. Thus, this study does not only reveal a novel nutraceutical ingredient, CCH-As, against liver fibrogenesis, but also offer an alternative way to expand the utilization of poultry byproducts.

Background Cardiac fibrosis (CF) plays a critical role in the pathogenesis of heart failure (HF), predisposing to contractile dysfunction, myocardial stiffness and life-threatening arrhythmias. Early detection and intervention of myocardial fibrosis, therefore, can delay or prevent the progression of HF. Current diagnostic and treatment options for CF, however, are very limited. Purpose To test the hypothesis that circulating lncRNA expression signature can be a sensitive biomarker to diagnose and correlate with the extent of CF in human heart. Methods Plasma RNA samples from healthy control (n=23) and from patients with diffuse myocardial fibrosis documented by cardiac magnetic resonance (T1 mapping technique, n=16) were analyzed by RNA sequencing (RNASeq), followed by differential expression, gene ontology/pathway and receiver operating characteristic (ROC) curve analyses. Results RNASeq revealed that 439 mRNAs were differentially expressed in the plasma from patients with CF, compared to that from controls. Gene ontology analyses showed significant enrichment of these CF-linked genes in cytoskeleton organization, innate immune response, and MAPK-Erk signaling, cellular processes known to contribute to the development of CF. Further analyses showed that 84 plasma lncRNAs were dysregulated in patients with CF; 5 of the lncRNAs (ENSGehz746.10420265401.1, ENSGehz746.10420229124.6, ENSGehz746.10420281376.1, ENSGehz746.10420273275.1 and ENSGehz746.10420179818.13) showed concordant changes with their cis-mRNA (UBB, VIM, BCL2L1, PPP3R1 and PCBP1). Hierarchical clustering analyses demonstrated that the expression signature of plasma lncRNAs could distinguish patients with CF from control patients (classification accuracy 87%). The expression levels of plasma lncRNAs ENSGehz746.10420258017.1 and ENSGehz746.10420265401.1, in particular, were markedly downregulated in patients with myocardial fibrosis. ROC curve analyses showed that low expression levels of ENSGehz746.10420258017.1 and ENSGehz746.10420265401.1 were highly sensitive and specific to detect the presence of CF in human heart (AUC 0.9 for both lncRNAs, Figure). LncRNA Predicts Cardiac Fibrosis Conclusions The present study revealed a circulating transcriptome signature that reflects the pathogenesis of CF. The expression profiles of circulating lncRNAs discriminated between patients with and without interstitial myocardial fibrosis. LncRNAs ENSGehz746.10420258017.1 and ENSGehz746.10420265401.1, in particular, are novel biomarkers with potential to facilitate clinical diagnosis of myocardial fibrosis.

Elongator complexes are well known to be involved in a wide variety of cellular processes; however, their functions in mammalian oocytes have not been characterized. Here, we demonstrated in mice that specific deletion of one of the core subunits, Ikbkap/Elp1, in oocytes resulted in spindle defects and chromosome disorganization without affecting folliculogenesis. In accordance with these findings, we observed that Ikbkap mutant female mice are subfertile. Further analyses uncovered that kinetochore–microtubule attachments are severely compromised in Ikbkap-deficient oocytes. Moreover, we revealed that Ikbkap modulates the acetylation status of α-tubulin in oocytes, which may at least in part mediate the meiotic phenotypes described above by affecting microtubule dynamics and kinetochore function. Finally, we showed that embryos derived from Ikbkap-deficient oocytes exhibit an increased frequency of aneuploidy, digyny, progressive delays in preimplantation development, and severe degeneration before reaching the blastocyst stage. In summary, we identify Ikbkap as an important player in regulating oocyte meiosis by modulating tubulin acetylation for chromosome/spindle organization.

Reactive oxygen species (ROS) overproduction is highly related to some human chronic diseases. There are approximately 400 metric tons of chalazae produced yearly after the processing of the liquid-egg production, which are disposed of as waste. The objectives of this study were to look for the optimal production condition of antioxidant crude chalaza hydrolysates and evaluate the in vivo antioxidant capacity via a chronic alcohol consumption mouse model.Antioxidant crude chalaza hydrolysates (CCH-As) could be produced by protease A at 1:100 ratio (w/w) and 0.5 h hydrolytic period. After our analyses, CCH-As were rich in leucine, arginine, phenylalanine, valine, lysine and antioxidant dipeptides (anserine and carnosine), and the major molecular masses were lower than 15 kDa. Regarding protective effects of CCH-As against oxidative damage in alcoholic-liquid-diet-fed mice, alcohol-fed mice had lower (P0.05) liver antioxidant capacities, and higher (P0.05) liver lipid contents, serum lipid/liver damage indices and IL-1β/IL-6 values. CCH-A supplementation reversed (P0.05) liver antioxidant capacities and reduced (P0.05) serum/liver lipids in alcohol-fed mice, which may result from increased (P0.05) fecal lipid output, upregulated (P0.05) fatty acid β-oxidation and downregulated (P0.05) lipogenesis in the liver.Taken together, this CCH-A should benefit the liquid-egg industry, while also offering consumers a choice of healthy ingredients from animal sources. © 2018 Society of Chemical Industry.

The cardiovascular and liver protection of carnitine (CNT) in a high-fat diet was investigated. Male C57BL/6 mice were divided randomly into four groups: 1) CON: Control, 2) HFD: high-fat diet, 3) CNTL: HFD + 500 mg CNT/kg BW, and 4) CNTH: HFD + 1500 mg CNT/kg BW. After a 25-week experimental period, CNT supplementation reduced (p

Background Kisspeptin, encoded by the Kiss1 gene, has many forms including kisspeptin54, kisspeptin14, kisspeptin13, and kisspeptin10, and all these peptides have the same affinity to their receptor KISS1R encoded by the Kiss1r gene. The KISS1–KISS1R system was discovered in neurons, and many reports stress on their function in the brain. However, recent studies have shown that Kiss1 and Kiss1r are expressed in the testes. The goal of this study was to demonstrate the roles of Kiss1 and Kiss1r in testicular function, especially their steroidogenic activity. Methods Kisspeptin10 and the kisspeptin10 antagonist peptide234 were used to determine their effect on testosterone production. Moreover, expression of steroidogenic genes in mouse testes and their gonadosomatic index (weight of the testes divided by the total body weight) and also serum testosterone level were studied between the ages of 2 weeks and 15 weeks. Results Kisspeptin10 and peptide234 did not affect testosterone production in primary Leydig cells from adult mice. Kiss1 and Esr1 expression also increased during puberty. The peak gonadosomatic index occurred at 4 weeks of age, and serum testosterone levels plateaued after the age of 4 weeks. Conclusion Our results suggest that kisspeptin10 does not affect steroidogenesis in adult Leydig cells, but its pattern of expression follows the stages of testicular development. Future studies should determine if kisspeptin regulates testicular development during puberty.

In this study, we identified novel family- and genus-specific DNA markers in Mugilidae fish. Genomic DNA was isolated from the blood of fish of 15 families and eighty (80) random primers were used for random amplified polymorphic DNA (RAPD) fingerprinting. When the primer OPAV04 was employed, a novel specific PCR product was observed in the Mugilidae family. In addition, another novel specific PCR product was also observed in the Liza genus while using primer OPAV10. Sequencing analysis revealed that the novel family- and genus-specific DNA fragments were 857 and 419 bp, respectively, and no similar sequences were found in GenBank. Two primers sets were designed based on the family- and genus-specific sequences to confirm the RAPD results and the 571 and 187 bp predicted bands were successfully amplified by PCR. Intriguingly, these two novel specific DNA markers were also effectively used for terrestrial and aquatic animal discrimination. Therefore, the novel family- and genus-specific DNA markers identified in this study can be used as an effective tool for rapid and accurate determination of the Mugilidae family and Liza genus, and even for cross-species identification. Key words: Mugilidae, family- and genus-specific sequences, DNA markers.

Nonalcoholic fatty liver (NAFL) is also called hepatic steatosis and has become an emergent liver disease in developed and developing nations. This study was to exam the preventive effects of taurine (Tau) on the development of hepatic steatosis via a hamster model. Although hepatic steatosis of hamsters was induced by feeding a high-fat/cholesterol diet, drinking water containing 0.35 and 0.7% Tau improved (p < 0.05) the serum lipid profile. Meanwhile, the smaller (p < 0.05) liver sizes and lower (p < 0.05) hepatic lipids in high-fat/cholesterol dietary hamsters drinking Tau may be partially due to higher (p < 0.05) fecal cholesterol, triacylglycerol, and bile acid outputs. In the regulation of lipid homeostasis, drinking a Tau solution upregulated (p < 0.05) low-density lipoprotein receptor and CYP7A1 gene expressions in high-fat/cholesterol dietary hamsters, which result in increased fecal cholesterol and bile acid outputs. Drinking a Tau solution also upregulated (p < 0.05) peroxisome proliferator-activated receptor-α (PPAR-α) and uncoupling protein 2 (UPC2) gene expressions in high-fat/cholesterol dietary hamsters, thus increasing energy expenditure. Besides, Tau also enhanced (p < 0.05) liver antioxidant capacities (GSH, TEAC, SOD, and CAT) and decreased (p < 0.05) lipid peroxidation (MDA), which alleviated liver damage in the high-fat/cholesterol dietary hamsters. Therefore, Tau shows preventive effects on the development of hepatic steatosis induced by a high-fat/cholesterol dietary habit.

We report for the first time the subcellular localization of endogenous Aurora-C and examine its roles during female mouse meiosis. The most dramatic effect observed in the oocyte injected with kinase-deficient Aurora-C mRNA is cytokinesis failure in meiosis I, resulting in production of large polyploid oocytes., We previously isolated Aurora-C/Aie1 in a screen for kinases expressed in mouse sperm and eggs. Here, we show the localization of endogenous Aurora-C and examine its roles during female mouse meiosis. Aurora-C was detected at the centromeres and along the chromosome arms in prometaphase I–metaphase I and was concentrated at centromeres at metaphase II, in which Aurora-C also was phosphorylated at Thr171. During the anaphase I–telophase I transition, Aurora-C was dephosphorylated and relocalized to the midzone and midbody. Microinjection of the kinase-deficient Aurora-C (AurC-KD) mRNA into mouse oocytes significantly inhibited Aurora-C activity and caused multiple defects, including chromosome misalignment, abnormal kinetochore–microtubule attachment, premature chromosome segregation, and cytokinesis failure in meiosis I. Furthermore, AurC-KD reduced Aurora-C and histone H3 phosphorylation and inhibited kinetochore localization of Bub1 and BubR1. Similar effects also were observed in the oocytes injected with INCNEP-delIN mRNAs, in which the Aurora-C binding motif was removed. The most dramatic effect observed in AurC-KD–injected oocytes is cytokinesis failure in meiosis I, resulting in producing large polyploid oocytes, a pattern similar to Aurora-C deficiency human spermatozoa. Surprisingly, we detected no Aurora-B protein in mouse oocytes. We propose that Aurora-C, but not Aurora-B, plays essential roles in female mouse meiosis.

Eight male hamsters per group were assigned randomly to one of the following diets: chow diet (Control); high-fat/cholesterol diet (HFCD); HFCD supplemented with 1% Tau (HFCD/1% Tau); HFCD supplemented with 2% Tau (HFCD/2% Tau). Tau supplementation improved (P 0.05) changed by Tau supplementation, faecal cholesterol and bile acid outputs were increased (P

A cDNA microarray-assisted experiment was conducted to survey genes that respond early to heat shock in enriched immature porcine germ cells; the 5′-UTR flanking the highest upregulated gene, heat shock 105/110 kDa protein 1 ( Hsph1 or Hsp105 ), in response to heat shock was also investigated. We established a porcine testis cDNA microarray with 9944 transcripts from two libraries constructed from the testes of mature boars, with or without heat shock. After a mild heat shock treatment (39 °C for 1 h and recovered at 34 °C for 2 h), 380 transcripts demonstrated significant gene expression in enriched immature germ cells; 326 were upregulated and 54 were downregulated. Ten transcripts of interest exhibiting significance analysis of microarrays (SAM) scores higher than the median were subjected to quantitative real-time PCR; three ( Hsp105 , Hspa4l and Thap4 ) were upregulated >1.5-fold. The sequence of the 5′-UTR of Hsp105 , the highest upregulated transcript, was cloned and analyzed. A single nucleotide polymorphism (SNP) was found at position −762 (C or T) upstream of the translational start site (ATG codon). Only two genotypes (CC or TC) were found in the mature boars that were studied ( n = 31). A heterozygous genotype (TC) at this SNP site revealed an elevated percentage of morphologically normal sperm during hot and cold seasons; this SNP may be a useful marker for semen quality in boars. Furthermore, the cell-model established from enriched primitive germ cells has potential for the study of reproduction in mature animals.

The purpose of this study was to characterize differentially expressed transcripts associated with varying rates of egg production in Taiwan country chickens. Ovarian follicles were isolated from two strains of chicken which showed low (B) or high (L2) rates of egg production, then processed for RNA extraction and cDNA library construction. Three thousand and eight forty clones were randomly selected from the cDNA library and amplified by PCR, then used in microarray analysis. Differentially expressed transcripts (P0.05, log(2)or = 1.75) were sequenced, and aligned using GenBank. This analysis revealed 20 non-redundant sequences which corresponded to known transcripts. Eight transcripts were expressed at a higher level in ovarian tissue prepared from chicken strain B, and 12 transcripts were expressed at a higher level in L2 birds. These differential patterns of expression were confirmed by semi-quantitative RT-PCR. We show that transcripts of cyclin B2 (cycB2), ferritin heavy polypeptide 1 (FTH1), Gag-Pol polyprotein, thymosin beta4 (TB4) and elongation factor 1 alpha1 (EEF1A1) were enriched in B strain ovarian follicles. In contrast, thioredoxin (TXN), acetyl-CoA dehydrogenase long chain (ACADL), inhibitor of growth family member 4 (ING4) and annexin II (ANXA2) were expressed in at higher levels in the L2 strain. We suggest that our approach may lead to the isolation of effective molecular markers that can be used in selection programs in Taiwan country chickens.

We have constructed a tissue-specific in-house cDNA microarray to identify differentially expressed transcripts in shell glands from low (B) and high (L2) egg production strains of Taiwanese country chickens during their egg-laying period. The shell gland cDNA library was constructed from the high egg production strain. cDNA clones (7680) were randomly selected and their 5′-end sequences characterized. After excluding overlapping sequences, an in-house cDNA microarray, representing 2743 non-redundant transcripts, was generated for functional genomic studies. Using our microarray, we have successfully identified 85 differentially expressed transcripts from the two different strains of chicken shell glands. In this study, 34 of these transcripts were associated with signal transduction, protein biosynthesis, cell adhesion, cellular metabolism, skeletal development, cell organization and biogenesis. We selected a number of the differentially expressed transcripts for further validation using semi-quantitative RT-PCR. These included elongation factor 2 (EEF2), ovocalyxin-32 (OCX-32) and annexin A2 (ANXA2) which were expressed at high levels in the chicken shell glands of the B strain and, in contrast, the coactosin-like protein (COTL1), transcription factor SOX18 and MX protein were more highly expressed in the L2 strain. Our results suggest that these differentially expressed transcripts may be suitable to use as molecular markers for high rates of egg production, and now need to be investigated further to assess whether they can be applied for use in breeding selection programs in Taiwanese country chickens.

That most Columbidae birds have no conspicuous sexual dimorphism often makes it difficult to identify their sex on the basis of external morphology. In the present study, we report a novel sex-specific DNA marker in Columbidae birds. DNA was extracted from one member of this bird group, Streptopelia orientalis (S. orientalis, oriental turtle dove), and used to identify a female-specific DNA marker using a random amplified polymorphic DNA (RAPD) fingerprinting. One hundred and sixty random primers were used for the RAPD-PCR reactions. When using the OPAV17 primer, a novel 902 bp sex-specific PCR product was amplified from known female birds. This fragment of DNA was cloned and sequenced. Two primers, TurSexOPAV17-F and TurSexOPAV17-R, were designed from the cloned sex-specific sequence, and were successfully used to amplify a 777 bp female-specific fragment using PCR from S. orientalis DNA. This sex-specific marker was also amplified from genomic DNA samples of two other female Columbidae, S. chinensis and Columba livia. Sequence analysis showed that this novel sex-specific marker was highly conserved amongst these three bird species. In contrast, the PCR product was not amplified from male DNA of these species, nor from either sex of the S. chinensis formosa birds. Therefore, we concluded that our novel marker can be used to rapidly and accurately identify the sex of birds from three species of Columbidae.

The meiotic generation of haploid gametes with equal contents of genetic material is important for sexual reproduction in mammals. Errors in the transmission of chromosomes during meiosis may lead to aneuploidy, which is the leading cause of miscarriage and congenital birth defects in humans. The Aurora kinases, which include Aurora-A, Aurora-B, and Aurora-C, are highly conserved serine–threonine kinases that play essential roles in centrosome function, chromosome segregation, and cytokinesis during mitosis and meiosis. While Aurora-A and Aurora-B have been extensively studied in mitosis, the role of Aurora-C in meiosis is only now starting to be revealed. For example, the perturbation of Aurora-C kinase activity by microinjection of Aurora-C-kinase-dead mutant mRNAs into mouse oocytes induced multiple defects, including chromosome misalignment, abnormal kinetochore–microtubule attachment, premature chromosome segregation, and failure of cytokinesis during meiotic division. However, the analysis of such defects is complicated by the possibility that Aurora-B may be present in mammalian germ cells. Interestingly, a homozygous mutation of Aurora-C in humans leads to the production of large-headed polyploid spermatozoa and causes male infertility, but homozygous females are fertile. Mouse studies regarding the roles of Aurora-B and Aurora-C in female meiotic divisions have yielded inconsistent results, and it has proven difficult to explain why homozygous human females have no significant clinical phenotype. In this review, we will discuss the controversial status of Aurora-B in oocytes and the possible role of Aurora-C during meiotic division.

We previously isolated Aurora-C (Aurkc/Aie1) in a screen for kinases expressed in mouse sperm and eggs. Aurora-C kinase was reported to be a chromosomal passenger protein that plays critical roles in chromosome alignment, segregation, kinetochore-microtubule attachment, and cytokinesis in female mouse meiosis. This chapter describes experimental approaches for examining the subcellular localization and function of Aurora-C kinase during female mouse meiosis, presenting detailed methods for introducing exogenous Aurora-C wild-type and kinase-dead mutant mRNAs into mouse oocytes by cytosolic microinjection, and preparing whole-mount meiotic oocytes and chromosome spreads for confocal immunofluorescence microscopy.

We previously isolated Aurora-C (Aurkc/Aie1) in a screen for kinases expressed in mouse sperm and eggs. Aurora-C kinase was reported to be a chromosomal passenger protein that plays critical roles in chromosome alignment, segregation, kinetochore-microtubule attachment, and cytokinesis in female mouse meiosis. This chapter describes experimental approaches for examining the subcellular localization and function of Aurora-C kinase during female mouse meiosis, presenting detailed methods for introducing exogenous Aurora-C wild-type and kinase-dead mutant mRNAs into mouse oocytes by cytosolic microinjection, and preparing whole-mount meiotic oocytes and chromosome spreads for confocal immunofluorescence microscopy.

We performed the first genome-wide expression analysis to compare the differences in gene expression in the female sperm reservoir of the duck reproductive tract between two groups with long and short fertile periods to identify factors that may be associated with the fertile period using an oligonucleotide microarray. RNA was extracted from the uterovaginal junction (UV junction) of the two groups. Affymetrix chips containing comprehensive coverage of 32773 transcripts were hybridized with biotin-labeled cRNA, and three biological repeats were performed. We identified 27 transcripts as being differentially regulated. Interestingly, by mapping the differentially expressed transcripts to annotated pathways, we found that Neuropeptide Y (NPY), the RNA expression of which was increased by 2.96-fold in the short-fertile-period group as compared with the long-fertile-period group in our experiment, has been shown to reduce blood flow and substance supply to local tissues. Enah/Vasp-like (EVL), the RNA expression of which was significantly increased by 1.77-fold in the short-fertile-period group as compared with the long-period group, has been demonstrated to be important in activated T-cells. In contrast, trafficking kinesin-binding protein 1 (TRAK1), the expression of which was increased by 2.33-fold in the long-period group as compared with its counterparts, has been suggested to inhibit precocious activation of sperm and prolong sperm life in the female sperm reservoir. The results of real-time PCR confirmed the data obtained by microarray analysis. Our study demonstrated that combining global gene expression investigation with annotated pathway resources contributes to the understanding of sperm life when sustained in the UV junction.

Cardiovascular protection of deep-seawater (DSW) drinking water was assessed using high-fat/cholesterol-fed hamsters in this study. All hamsters were fed a high-fat/cholesterol diet (12% fat/0.2% cholesterol), and drinking solutions were normal distiled water (NDW, hardness: 2.48 ppm), DSW300 (hardness: 324.5 ppm), DSW900 (hardness: 858.5 ppm), and DSW1500 (hardness: 1569.0 ppm), respectively. After a 6-week feeding period, body weight, heart rates, and blood pressures of hamsters were not influenced by DSW drinking waters. Serum total cholesterol (TC), triacylglycerol (TAG), atherogenic index, and malondialdehyde (MDA) levels were decreased (p < 0.05) in the DSW-drinking-water groups, as compared to those in the NDW group. Additionally, increased (p < 0.05) serum Trolox equivalent antioxidant capacity (TEAC), and faecal TC, TAG, and bile acid outputs were measured in the DSW-drinking-water groups. Hepatic low-density-lipoprotein receptor (LDL receptor) and cholesterol-7α-hydroxylase (CYP7A1) gene expressions were upregulated (p < 0.05) by DSW drinking waters. These results demonstrate that DSW drinking water benefits the attenuation of high-fat/cholesterol-diet-induced cardiovascular disorders in hamsters.

Amplified fragment length polymorphism (AFLP) with multicolored fluorescent molecular markers was used to analyze duck (Anas platyrhynchos) genomic DNA and to construct the first AFLP genetic linkage map. These markers were developed and genotyped in 766 F2 individuals from six families from a cross between two different selected duck lines, brown Tsaiya and Pekin. Two hundred and ninety-six polymorphic bands (64% of all bands) were detected using 18 pairs of fluorescent TaqI/EcoRI primer combinations. Each primer set produced a range of 7 to 29 fragments in the reactions, and generated on average 16.4 polymorphic bands. The AFLP linkage map included 260 co-dominant markers distributed in 32 linkage groups. Twenty-one co-dominant markers were not linked with any other marker. Each linkage group contained three to 63 molecular markers and their size ranged between 19.0 cM and 171.9 cM. This AFLP linkage map provides important information for establishing a duck chromosome map, for mapping quantitative trait loci (QTL mapping) and for breeding applications.

1. The accuracy and reproducibility of AFLP fingerprinting was investigated in the duck (Anas Platyrhynchos), using a multicolour fluorescent labeling technique. The fluorescent labelling fragments were separated on a capillary electrophoresis-base ABI PRISM 3100 Genetic Analyzer. 2. A total of 337 AFLP peaks with 103 of them being polymorphic markers were generated by 16 sets consisting of EcoRI/TaqI primer pair combinations. The number and size range of AFLP polymorphisms detected per primer pair varied from 3 to 11 and 58 to 290 bp, respectively. About 30.6% (103/337) of AFLP peaks were detected polymorphisms, with an average of 6.4 polymorphic markers per primer pair. 3. The clear polymorphic peaks were amplified with EcoR+AC/Taq+AC primer combinations. The AFLP peaks showed high reproducibility. From the family testing, we found that the fingerprints of all the offspring were derived from one or other parent. Therefore, we conclude that AFLP fingerprinting might be a suitable method for duck paternity testing.

Ostrich absence of heteromorphic sex chromosomes, unique sequences or markers located in the ostrich W-chromosome. Random amplified polymorphic DNA (RAPD) fingerprinting was carried out to investigate the sex-specific DNA sequence for sexing in ostrich. One hundred and forty random primers were used for random amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR). One of these primers, OPAJ-13, produced a sex-specific band only found in tested females, which was isolated and constructed into plasmids for nucleotide sequencing. A 760 bp novel female-specific sequence was obtained. Two primers (OstSexOPAJ13-F and -R) were designed according to the cloned female sequence to amplify the female-specific fragment from genomic DNA of ostriches for sexing by PCR. The sex-specific band was represented in females but none were found in the males. This result showed that the sex of ostrich could be easily and effectively identified using the female-specific primers for PCR technique.