Hybridization chain reaction (HCR) is a kind of self-assembly reaction which can be carried out without enzyme participation at room temperature. It is a new type of signal amplification method. It is not only widely used in the common areas, such as colorimetric, fluorescent and electrochemical method, but also in the new field of Chemiluminescence, Electrochemical luminescence, Photochemiluminescence and Raman spectra and so on. So biosensor based on HCR can be used to detect nucleic acids, proteins, cells, small molecules and other biological molecules. This paper reviews the application of the HCR-based biosensors in the biosensing field in the nearly four years, and the future development trend is prospected.

Circular RNA (circRNA) is a group of non-coding RNA (ncRNA) characterized by the presence of a covalent bond linking 3' and 5' ends produced by backsplicing. It widely spreads and has the features of construct stability, sequence conservative, cell or tissue-specific expression and so on. So far, circRNA has been found exist in different species extensively and already three classes of circRNA have been detected, including exonic circRNA, circular intronic RNA (ciRNA) and exon-intron RNA (ElciRNA). Concerning the biogenesis of these circRNAs, scientists purposed six models in total, among them, three models clarified the formation of exonic circRNA and ElciRNA, and the others explained the generation of ciRNA. As research continues, there were lots of experiments support these models directly or indirectly. Moreover, some papers reported that the biogenesis of circRNA was regulated by splicing factors, such as SR, hnRNP and RBM20 etc. Relative researches showed that circRNA performs its function through various channels. Part of circRNAs act as natural microRNA (miRNA) spong to regulate the expression of them, and ciRNA or ElciRNA interacts with snRNP and recruits RNA Polymerase Ⅱ in the promoter region of the host-gene around where they are derived to regulate gene expression in cis. Besides, some circRNAs could regulate cell cycle or ageing etc. by interacting with protein. Although there was no evidence to show that circRNA can be translated in eukaryotes, some groups have found a number of virus circRNAs and viroid circRNAs could be translated into protein. Notably, circRNA is closely related to the diseases such as cancer (including breast cancer, lung cancer, hepatocellular carcinoma etc.) and cardiopathy, which are known as human healthy killer, it also plays a role in neurodevelopment, immune and other physiological process, indicating its significant value. This paper summarized the formation mechanism and features of circRNA, as well as the recent advance of circRNA in mainly function, relative research methods and the relationship with disease, at the last part of this paper, we discussed the unresolved issues about circRNA, such as which type of spliceosomes participate in back-splicing and how do them assembly and work? How the nucleocytoplasmic export of circRNA is regulated? as well as how circRNA is ultimately degraded. In general, circRNA extend the diversity and complexity of eukaryotic transcriptomes, and we believe that delve into circRNA will enrich the study content of the post-genome era and provide theoretical basis and technical guidance for better understanding, diagnosing and treatment of human diseases

Thyriod hormone responsive spot 14 (THRSP) is a transcriptional regulation factor which participates in regulating the activity of lipogenic enzymes and in turn affects the lipid metabolism in animal tissues. It includes α and β two subtypes. In this study, in order to investigate the effects of THRSPα on the carcass and abdominal fat traits and explore its expression profile in Jinghai Yellow chicken (Gallus gallus), polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) combining sequencing method was used to detect the polymorphisms of THRSPα , and qRT-PCR technology was applied to analyze the expression level of this gene in 10 tissues (including heart, liver, spleen, lung, kidney, breast muscle, leg muscle, abdominal fat, small intestine and hypothalamus) and the expression profile of this gene from 0 to 16 weeks of age in Jinghai Yellow chicken. As a result, 2 mutations (C129T and T160G) were detected in exon 2 of THRSPα. Three haplotypes and 5 diplotypes were formed in the group of Jinghai Yellow chicken. Association analysis showed that the polymorphisms of THRSPα were significantly associated with the abdominal fat weight of Jinghai Yellow chicken (P＜0.01). Multiple comparisons between different diplotypes showed that no significant differences (P＞0.05) were found in the live weight, breast muscle weight, leg muscle weight, eviscerated weight and half eviscerated weight of Jinghai Yellow chicken. However, significant differences at the level of P＜0.05 and P＜0.01 were found in the carcass weight and abdominal fat weight, respectively. In the aspect of carcass weight, H1H3 diplotype has the highest value which was significantly higher than H1H1 and H2H2 diplotypes (P＜0.05), while no significant differences were found between H1H1, H1H2 and H2H2 (P＞0.05). In terms of abdominal fat weight, H1H3 also has the highest abdominal fat weight which was significantly higher than H1H1 and H2H2 at the level of P＜0.01. The abdominal fat weight of H2H2 was the lowest which was significantly lower than any other diplotypes (P＜0.01). The results of the expression level of THRSPα in tissues revealed that the expression level of THRSPα was higher in liver and abdominal fat compared to almost no expression in heart, spleen, lung, kidney, breast muscle, leg muscle, small intestine and hypothalamus. According to the results of the expression profile, the expression of THRSPα in liver had a trend of rise first then fall with the highest expression level at 8 weeks of age and the lowest expression level at 0 week of age. However, the expression level in abdominal fat tissue was relatively stable. Comparing the expression level in these two tissues at different weeks of age, the expression level of THRSPα in liver was much higher than the expression level in abdominal fat from 4 to 12 weeks of age, while at 0 and 16 weeks of age, the expression level in liver was lower than in abdominal fat tissue. Therefore, these results further demonstrated that THRSPα was the candidate gene of chicken's carcass and abdominal fat traits. And the regulatory mechanism of THRSPα was revealed preliminarily on the fat synthesis and deposition in liver and abdominal fat.

Complement 9 is involved in the membrane attack complex (MAC) forming with other complement molecules on the target cell membrane, and takes part in the immunoreaction through the target cell disruption mediated by MAC. Single nucleotide polymorphisms (SNPs) analysis of genes functionally related to the streptococcosis resistance of nile tilapia (Oreochromis niloticus) can provide useful information for tilapia molecular breeding. In this study, the genomic fragments of complement 9 gene in O. niloticus (OnC9) were amplified, which contain 11 exons and 10 introns. Then 22 SNPs were identified by using directly sequencing and blasting. The SNPs S1 (C483T), S3 (G954A), S4 (T1180C), S5 (G1539A), S6 (G1627T), S7 (A2590T), S10 (T2876G), S11 (C2915G), S12 (T2939G), S13 (A2942C), S14 (A3544C), S15 (T3604C), S16 (T3670C), S17 (T3841C), S18 (T4339C), S19 (G4494C), S20 (A4678G), S21 (T5790A), and S22 (T5816G) were in introns of OnC9, while the SNPs S2 (C644T), S8 (C2660T), and S9 (A2751T) were in exons. Half of the SNPs in OnC9 was produced by base transition, and half of them was produced by base transversion. In addition, S8 was synonymous mutation and S2, S9 belong to the non-synonymous mutations. 81 tilapias from the resistance group (RG) and 84 tilapias from the susceptible group (SG) were used to analyze genotype polymorphisms by snapshot method. The polymorphisms and genetic parameter of the OnC9 SNPs in SG and RG were calculated by the software Popgen32. The result showed that the polymorphism information content (PIC) of the OnC9 SNPs was 0.09~0.37, suggesting that all the SNPs locus had moderate or intermediate polymorphism. In order to analyze the correlation between the OnC9 SNPs and the phenotype of streptococcosis resistance, the genotype frequency, allele frequency of the 22 SNPs in OnC9 was analyzed by SPSS 20 and tested by chi-square test. The association analysis showed that Two SNPs (S8 and S21) were significantly associated with streptococcosis resistance (P＜0.05). Moreover, all the OnC9 SNPs could formed 4 haplotype blocks and 11 haplotypes from the prediction of haplotypes and linkage disequilibrium analysis used software Haploview 4.2. And 2 of the haplotypes (GTTAG and ATGA) were significantly associated with Streptococcus agalactiae susceptible (P＜0.05). These results indicate that the OnC9 gene can be a candidate for tilapia Streptococcus agalactiae resistance traits which could be useful for molecular breeding of O.niloticus.

Coix (Coix lachryma-jobi) is a crop, which has high value of nutrition and medicine. However, the most studies are focus on physiology and biochemistry, there is a lack of molecular biology research in coix. In order to obtain genetic datas and functions, the study carried on the transcriptome analysis of coix leaf in seedlings. In this study, 92 865 unigenes were obtained and 35 813 unigenes were successfully annotated to some databases, being 39.87% of total unigenes numbers. GO (Gene Ontology) analysis showed that 1 053 unigenes were identi?ed as potentially involved in the drought, which 409 and 125 unigenes were identified as encoding some plant hormones and osmosis synthetases. Meanwhile, some genes and enzymes were also annotated in GO database, including lipid compounds, sterol and catechol. For KEGG (Kyoto Encyclopedia of Genes and Genomes) analysis, 6 560 unigenes were assigned to 170 known pathways. Among these pathways, "Plant hormone signal transduction" (264 DEGs), which were the more enriched metabolic pathways of differentially expressed genes. Among plant hormone signal transduction, AMPK (AMP-activated protein kinase), MAPK (mitogen-activated protein kinase) and Ca signal transduction were related to drought stress. In addition, 7 534 simple sequence repeats were detected. The most abundant microsatellites were mononucleotide repeats, with the SSR numbers being 3 068, followed by trinucleotide and double base repeats, being 3 017 and 1 288, respectively. This study will provide the foundation and reference for functional genes obtaining and molecular biology study in coix.

Phenol is a key chemical engineering material, which is detrimental to the agroecological environment. In Thauera aromatica strain K172, a denitrifying bacterium, phenol undergoes phosphorylation and carboxylation and is converted into 4-hydroxybenzoic acid, which is less toxic. Phenylphosphate carboxylase, consists of α, δ, β and γ subunits, is a key enzyme of phenol degradation process. This study used the open reading frames, orf 5 and orf 12, which encode the δ and γ subunits, as the templates of DNA probes to analyze the diversity of related genes in strains of T. aromatica and Azoarcus evansii by Southern blot. The results indicated that these strains all contained several orf 5 homologs. The hybridization map showed there was no genus specificity of the gene. By contrast, orf 12 was only present in T. aromatica K172, S100 and A. evansii T and the gene cluster related to phenol degradation had a single copy in the genome. The gene was mapped to a PstⅠ digest fragment whose size was consistent among the bacterial strains. While in the bacterial culture experiments, only these 3 strains grew steadily on phenol as the only carbon source under the condition of denitrification. Therefore, our results suggested that T. aromatica K172, S100 and A. evansii T possessed the ability of phenol degradation through the same pathway. It is possible that the bacteria acquired the identical genes or gene cluster during evolution. This study provids new evidence of previous research that γ protein encoded by orf 12 is one subunit of phenylphosphate carboxylase. Meanwhile this work offers available scientific evidences for further studies of phenol degradation.

Mycoplasmal pneumonia of swine (MPS, also referred to as 'enzootic pneumonia', EP) caused by Mycoplasma hyopneumoniae (Mhp), is a chronic respiratory disease with the clinical sign of persistent dry cough. Epidemic of MPS has a negative impact on growth rate and feed conversion. The causative agent Mhp is easily infected and it will become worse with co-infections, which always gives rise to huge economic damage to the swine (Sus scrofa) industry, both at home and abroad. At present, most research is centered on pathological diagnosis, immunity, prevention and control. However, these medical treatments and public health measures are incapable of alleviating the negative impacts from MPS. For better solving the occurrence of MPS, we intend to adopt bioinformatics analysis methods to explore MPS from the molecular level for better illustrating its mechanism. For the purpose, this study was mainly based on Gene Ontology (GO) enrichment analysis, KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway analysis, text mining and other bioinformatics tools to complete the PigQTLdb (pig QTL Database) retrieval and biological annotations for MPS candidate genes. The results showed that 47 possible QTLs were associated with MPS with the full use of PigQTLdb, 2 of which were related to MPS susceptibility, and the others were involved in immunity. 38 GO terms linked to MPS were searched by CoPub, a text-mining tool, and then 3 379 genes contained in 47 QTLs were effective after semantic similarities calculation with R programming and GOSim package. Among them, 36 genes were statistically significant. For another, there were 53 genes found in GO enrichment analysis, and 16 genes in 'Asthma' pathway from KEGG analysis. In addition, six candidate genes were from the results of text-mining, including TNFα (tumor necrosis factor), NRAMP1 (natural resistance-associated macrophage protein 1), TCRs (T-cell receptor genes), CYP1A1 (cytochrome P450 family 1 subfamily A member 1), CYP3A29 (cytochrome P450 3A29v4) and SERPING1 (serpin family G member 1). As a result, there were totally 104 optimized genes for the MPS candidate gene set. Special attention should be paid on gene TNF, for it was involved in both GO enrichment analysis and KEGG pathway analysis results, and appeared in the text mining results as well. To be specific, TNF was one of the crucial factors to induce inflammation, which ranked fourth in significance among all candidate genes during calculations. We inferred that TNF and POU5F1 were the candidate genes of vital importance during MPS infections. These findings will offer a new aspect for the further research in the future, though there would be some improvements in the analysis, and these conclusions remained to be proved. In next step, we will make efforts to explain the relationship between MPS and other important economic traits and explore the molecular differences between Chinese indigenous pigs and western imported pigs for further to indicate its heritance mechanism and provide insights for human asthma study one day.

Transgenic herbicide-resistant soybean (Glycine max) is one of the most widely planted genetically modified crops in the world. Meanwhile, the impacts of its rapid extension brought to environmental have become a focus of public interests. In this study, the Korean quails (Coturnix japonica) were selected as test animals, which were closely related to the environment, were randomly divided into 5 treatment groups. To assess the safety of transgenic herbicide-resistant soybean to birds, a 90 d feeding trial with transgenic soybean (ZZ-J9331) and its non-transgenic counterpart (ZH-30) were supplied to quails in basal feed with a ratio of 28% and 70% respectively. After 90 d feeding, the influence of genetically modified (GM) soybean were evaluated by monitoring quails health and physiological indexes, recording animal's body weight and food intake, dissecting animals and calculating viscera coefficient afterwards. It was observed after 90 d, animals of each group were in good condition. Quails in each group ate normal, moved freely, and had shiny hair, nose, eyes and mouth without abnormal secretions. The body weight of all experimental groups showed an increasing trend, and no abnormal changes in food intake. Different from the previous literature, in addition to the above related to the growth and development index, this study collected blood samples from quail at the end of the experiment, blood and blood biochemical detection, in addition, this study also had some important organs pathological dyeing processing, in order to observe the possible toxicity of glyphosate tolerance of transgenic soybean on quails from the toxicological perspective. Compared with the non-GM soybean control group, several indexes of the serum biochemistry, hematology, relative organ weight of quails had statistically significant difference (P＜0.05) and no dose-relative or gender-relative, indicating no obvious biological significance. Moreover, no pathological change was observed in heart, brain, kidney, lung, stomach, intestine, spleen, adrenal gland, uterus, ovary, testis or epididymis, while fatty degeneration of hepatic cells and diffuse hyperplasia of fiber connective tissue were noticed in liver slices between experimental and control based diets group, which was taken as spontaneous lesions of liver, excluding the influence of absorbing transgenic soybean. Consequently, the transgenic herbicide-resistant soybean J9331 is as same safety as its non-GM maize in 90 d feeding study, which provides scientific evidence for the safe usage of GM soybean. In the aspects of the chronic toxicity of glyphosate herbicide resistant soybeans genetically (ZZ-J9331), this study fills the blank in the effects of transgenic soybean on bird potential environmental toxicology, provides data support for the commercial application, provide a scientific basis for environmental safety assessment and management of transgenic organisms in China.

Rhizobium type Ⅲ secretion system (T3SS) effector proteins play important roles in nodulation, nitrogen fixation and decision of host range. In order to express and analysis the type Ⅲ-like secretion system effector proteins of Azorhizobium caulinodans ORS571, the whole proteome of ORS571 was analyzed by bioinformatics software BPBAac and Effective T3 and related literature reports. Seven candidate effector proteins of ORS571 were obtained, and they were AZC_0308, AZC_0649, AZC_0667, AZC_2699, AZC_2928, AZC_3165 and AZC_4087, respectively. Specific amplification primers with restriction enzyme cutting sites were designed and synthesized, and whole gene sequences of the 7 effector proteins were obtained by PCR. Than the 7 gene fragments were cloned into the prokaryotic expression vector pMAL-c4x, respectively. Through optimizing the expression conditions, a large amount of soluble fusion proteins were obtained. Amylose Resin affinity chromatography was used to purify fusion proteins, and the results indicated that purification effect was better. Detection of fusion proteins expression were completed by Western blot using anti- maltose binding protein (MBP) monoclonal antibody (horse radish peroxidase conjugated), and the results implied that these 7 fusion proteins expressed correctly and the effect of protein expression was better. AZC_2928 gene deletion mutant was constructed and infected wheat (Triticum aestivum) XiaoYan 22, and the results showed that AZC_2928 gene had inhibitory effect on the growth of wheat. In conclusion, this research lays a foundation for the application of the Azorhizobium caulinodans ORS571 type Ⅲ-like secretion system effector proteins in symbiotic nitrogen fixation of non-leguminous plants.

Phosphoribosylpyrophosphate synthetases 2 gene (PRPS2) is the key enzyme of biological synthesis, which plays important role in controlling tumor development, promoting tumor cell's apoptosis, inhibiting cell proliferation and promoting spermatogenesis. In order to study the role of PRPS2 gene in pig (Sus scrofa) male sterility and the function of the protein encoded by the gene, using the PRPS2 mRNA sequences of pig and related species from GenBank as reference gene sequences, Specific primers were designed and Banna mini-pig inbred line (BMI) PRPS2 gene was amplified. qRT-PCR was applied to analysis the mRNA expression profiles of 15 important tissues. Protein sequence was used to carry out functional bioinformatics analysis. cDNA sequence of 1 282 bp (GenBank accession number: KU705637, the corresponding amino acid sequence accession number: AOC89055) of BMI PRPS2 was obtained, which encodes a protein of 318 amino acids. Tissue expression profile indicated that PRPS2 gene expressed highly in the seminal vesicle gland, and low expression in the other 14 tissues. Bioinformatics analysis indicated that PRPS2 protein molecular weight 34.83 kD, and isoelectric point 6.15, contained 2 conserved domains, no transmembrane region; no signal peptide sequences, its N-terminal and the C-terminal were hydrophilic; it had 4 types of functional active sites; the probability of being located in the cytoplasm was 89%; PRPS2 gene highly conserved in evolution. The results of the study will lay a foundation for further study of the gene about its functions in pig male sterility.

Pituitary adenylate cyclase-activating polypeptide (PACAP) is synthesized and secreted by the hypothalamus. As a bioactivator, it can stimulate the synthesis and secretion of growth hormone (GH) to promote the growth of animals. The objective of this study was to obtain full-length cDNA of pig (Sus scrofa) PACAP gene (pPACAP), and to analyse the relative expression level of PACAP gene during the differentiation of 3T3-L1 preadipocytes into adipocytes and then analyze the relative expression levels of PACAP, FAS, ACC mRNA and detect the target protein pacap-EGFP after the recombinant plasmid pEGFP-PACAP was transfected into 3T3-L1 cells in 48 h. Besides, cell counting kit-8 (CCK-8) was applied to see the growth curve of 3T3-L1 cells transfected. Firstly lipid droplet and the expression levels of PACAP gene were detected respectively by Oil red O and qRT-PCR after XDI was induced to differentiate the 3T3-L1 preadipocytes into adipocytes. And then primers with EcoRⅠand BglⅡ restriction enzymes were designed and amplified by PCR in accordance with the deposited sequence of pPACAP in GenBank. Further, the digested PCR product was ligated into eukaryotic vector pEGFP-C1 and the recombinant plasmid pEGFP-PACAP was successfully constructed. During the transfection in 48 h, the cell growth was observed with CCK-8 under in vitro transfection with XfectTM transfection reagent. Besides, the expected mRNA including PACAP, FAS and ACC and target protein pacap were successfully detected in 3T3-L1 cells, respectively by using qRT-PCR and Western blot. The adipogenic differentiation results showed that the expression level of PACAP of mRNA is gradually increasing and is the highest at 4 day after 3T3-L1 cells were induced; Double restriction enzymes digestion and sequence results indicated that the eukaryotic expression vector of pPACAP gene is successfully cloned, named as pEGFP-PACAP; By fluorescence microscopy, cells of the empty plasmid pEGFP-C1 transfection group and the recombinant plasmid pEGFP-PACAP transfection group both emit green fluorescence with high transfectection efficiency after 48 h post transfection. The growth curve showed that the cells grew well. Both mRNA and protein can expectedly express in 3T3-L1 cells including PACAP, FAS, ACC mRNA and pacap-EGFP protein. Especially the relative expression levels of PACAP, FAS, ACC mRNA in the pEGFP-PACAP group are significantly higher than the pEGFP-C1 group. All the results indicated PACAP may participate in adipogenesis. This study can be the theory basis for further study on adipogenesis and metabolism for pig in vitro.

Toll like receptor 1 (TLR1) is one of the important immune genes for cattle (Bos taurus), which has significant effects on immune recognition and inflammatory reponse. To investigate the correlation between the SNPs in the promoter region of TLR1 gene and somatic cell score (SCS), clinical mastitis (CM) and milking traits for Chinese Holstein cattle, the SNP in the promoter region of TLR1 gene was screened by PCR and direct sequencing for 20 cows with low SCS and 20 cows with high SCS. Finally, TLR1-245 G＞T for 866 Chinese Holstein cattle were detected using flight mass spectrometry. The SCS, CM and milking traits of tested cattles were collected from the management system of dairy farm. The correlation between the SNPs and these traits of tested cattles was analyzed using multi-factor variance analysis. The results showed that only one transcription factor binding site for TLR1-245 G＞T site (TATA binding protein, TBP) was found. The dominant genotype and gene of TLR1-245 were GT and T, and their frequencies were 0.487 and 0.584, respectively. The distribution of genotype was in the Hardy-Weinberg equilibrium. TLR1-245 G＞T showed extremely significant correlation with test-day milk yield (TDMY) and SCS (P＜0.01), and no significant correlation with fat content (FC), protein content (PC), lactose content (LC), total solid (TS) and milk urea nitrogen (MUN) (P＞0.05). The individuals cattles with TT genotype had higher TDMY than that of GG and GT, and individuals cattles with GG genotype had the lowest TDMY. The SCS for individuals cattles with GG genotype were significantly higher than that of GT and TT genotype (P＜0.05). TLR1-245 G＞T showed significant correlation with the number of cattle suffering from CM (P＜0.05), and the number of cattle suffering from CM for individuals with TT genotype was significantly lower than that of GG and GT genotype (P＜0.05). These results provide a reference for the increase of milk yield and the control of SCS in Chinese Holstein cattles.

Peste des petits ruminants (PPR) is a disease of major economic importance and imposes a significant constraint upon sheep(Ovis aries) and goat (Capra hircus) production owing to its high mortality rate. It is an acute, highly contagious and frequently fatal disease of sheep and goats caused by Peste des petits ruminants virus (PPRV), a member of genus Morbillivirus of family paramyxoviridae. In order to study the F gene sequence of PPRV GS strain and the immunogenicity of its prokaryotic expression products, the primers were designed according to the F gene sequence of PPRV Nigeria 75/1 strain in GenBank(GenBank No. HQ197753), and the F gene of PPRV GS strain was amplified by RT-PCR. Based on sequencing and sequence analysis, the primers were designed and the Fa fragment of F gene without the signal peptide and transmembrane domain was amplified. Then the recombinant plasmid pET-PPRV-Fa was constructed through cloning Fa fragment into pET-32a(+) and the recombinant plasmid was transformed into Escherichia coli Transetta (DE3). After induced by isopropyl β-D-1-thiogalactopyranoside (IPTG), the recombinant protein was expressed, and the New Zealand rabbit (Oryctolagus cuniculus) was immunized with purified protein, then the polyclonal antibody against PPRV fusion protein was prepared. Subsequently, the analysis of its immunogenicity was accomplished using indirect enzyme-linked immunosorbent assay (ELISA) and Western blot. The results showed that the complete CDS of F gene (GenBank No. KX822738) from PPRV GS strain was 1 641 bp encoding a 546 amino acids protein. The Fa fragment of F gene was successfully expressed in E. coli, whose molecular weights approximately was 59 kD and mainly existed in the form of inclusion body. The indirect ELISA and Western blot results showed that the prokaryotic expression products of Fa gene of PPRV GS strain had good immunogenicity. In conclusion, the research results provide theoretical basis for the study on the function of fusion protein from PPRV GS strain, and for developing a rapid diagnosis method.

The COR413 (cold-regulated) gene, which encodes a plant-specific cold responsive protein, plays a critical role in cold acclimation. To determine the functional characterization of COR413 genes in response to cold stress, integrated grapevine (Vitis vinifera) genomic approaches of data mining and homologous cloning were used to obtain a total of 4 COR413 gene family members from cold-tolerant Vitis amurensis, designated as VaCOR413-PM2A、-PM1X1、-IM1 and -PM2B, respectively. Sequence analysis revealed that the 4 COR413 family members from V. amurensis showed significant nucleotide (97.2%~100%) and amino acid (99.5%~100%) similarity with those from V. vinifera, and the nucleotide differences were several single nucleotide substitutions. All COR413 proteins from V. amurensis contained the conserved domain WCOR413, several transmembrane motifs and Serine/Threonine/Tyrosine phosphorylation sties, where a predicted N-terminal signal peptide was only found in VaCOR413-PM1X1 and a glycosylphosphatidylinositol (GPI) anchor was only observed in VaCOR413-PM2B. The 4 members of COR413 from 2 grapevine species can be grouped into 2 distinct clusters, where COR413-PM2A, -PM1X1 and -PM2B were grouped together and -IM1 fall into another cluster. Moreover, tissue-specific expression profile of COR413 gene family members and their responses to cold stress were analyzed in both V. amurensis and V. vinifera cv. Cabernet Sauvignon. The results showed that VaCOR413-PM2A, -PM1X1, -IM1 and -PM2B displayed different tissue-specific features that were also not similar to those from V. vinifera. Compared to the corresponding gene expression profiles in V. vinifera, VaCOR413-PM2A, -PM1X1, -IM1 and -PM2B were detected in all tissues of V. amurensis with different expression intensities, among which VaCOR413-PM2A and -PM1X1 showed a relatively high expression in shoots, tendrils, flowers and young leaves, whereas no signals of VvCOR413-PM1X1 and -IM1 were detected in V. vinifera. Analysis of the gene expression profiles during the time-course of cold stress demonstrated the existence of 2 groups with different expression profiles in V. amurensis and 3 groups in V. vinifera. In one group of V. amurensis containing VaCOR413-PM2A, -PM2B and -PM1X1, a similar expression pattern was observed in with a decreased tendency following the time-course of cold stress, while VaCOR413-IM1 displayed a distinct expression profiles from those in group Ⅰ, and a high level of VaCOR413-IM1 expression was detected 24 h after cold stress with 9.57-fold than that of unstressed control. In V. vinifera, the expression profile of VvCOR413-PM2A was similar to VvCOR413-PM2B and their gene induction reached a maximum at 3 h in response to cold stress. VvCOR413-PM1X1 was rapid and transient in response to cold treatment, reached a maximum at 1 h, and then decreased. VvCOR413-IM1 displayed a high level of expression at 3 h of cold stress with 3.65-fold than that of control, quickly decreased at 6 h and maintained a relatively low level to the cold stress. Data from cold stress treatment suggested that VaCOR413-PM2A, -PM1X1 and -PM2B which were grouped together, displayed down-regulated expression in response to cold stress and the degree of expression level of these genes were generally lower than those from V. vinifera. Interestingly, VaCOR413-IM1 categorized into another group, showed a significant induction feature 24 h after cold stress. The results provide reference basis for exploring the possible function and mechanism of the members of the family in response to cold stress, which lays the theoretical for cold-(frozen-) tolerant molecular improvement in grapevine.

The horse (Equus caballus) temperament traits directly determines whether or how it is trained. Mongolian horse is a natural species, mixes with fewer external genome, which makes it as the most suitable species for researching the association of the polymorphism of candidate genes with the phenotype of temperament traits. The aim of this study was to identify the polymorphisms of the solute carrier family 6 member 4 gene (SLC6A4), dopamine beta-hydroxylase gene (DBH), brain derived neurotrophic factor gene (BDNF) and catechol-O-methyltransferase gene(COMT) and provide insight into the association with the Mongolian horse temperament behavior. The research animals were consisted with 60 Mongolian horses, with the same breeding and training situation, which were numbered and collected peripheral blood, respectively. The 28 temperament behavioral characteristics were described with the questionnaire survey method. The SNP analysis for 4 candidate genes was performed using the single strand conformational polymorphism (SSCP) and gene sequencing. Further, the statistical analysis system (SAS) software was used to analyze the genotype and the association. For BDNF gene, the SNPs in 222 bp (C→T) and 261 bp (G→C) were identified, and the association analysis showed that the biting behavior was significant affected by BDNF genes as C222T and G261C. For DBH gene, the SNP was identified in 758 bp (G→A), and DBH gene (G758A) was significant associated with endurance and on other horse-friendly behavior, which also had remarkable influence on such behaviors as comprehension ability, crossing the river and swimming, sand bath et al. For SLC6A4 gene, the SNP was identified in 1 362 bp (T→C), and gene (T1362C) significantly affected the behavior such as nervousness, panic, the reaction to harness and timid, et al. SLC6A4 gene (T1362C) was remarkable associated with the independence acts else. For COMT gene, the SNP was identified in 217 bp (G→A) and COMT gene (G217A) had a greatly significant associated with the competitiveness. Above information illustrated that BDNF, DBH, SLC6A4 and COMT were associated with the Mongolian horses' behavior trait. This study can provide the theory basis for the Mongolian horse temperament trait's marker assisted selection.

Sugarcane (Saccharum species hybrid) is the most important crop for the production of sugar. Due to its high biomass production and lengthy growing season, sugarcane needs to absorb a large amount of potassium (K) throughout its life cycle. In the southern China where sugarcane is cultivated, most of the soil have low content of available K, which limits the production of sugarcane. To improve the productivity, one of the efficient means is to improve sugarcane cultivars that have efficient uptake capacity of K and hence are tolerant to low K. This study was conducted in which a major commercial cultivar, ROC22, was subject to low K stress. From the experiment, a potassium transporter gene, named SsHAK2 (GenBank accession number: KM98738), was cloned by reverse transcription-polymerase chainreaction (RT-PCR) from total RNA of sugarcane roots. The full length of the SsHAK2 gene was 2 798 bp, and contained a complete open reading frame of 2 352 bp encoding a protein with 784 amino acids. The molecular weight of SsHAK2 was 87.602 kD and the isoelectric point of 8.85, as the alkaline protein. It contained twelve transmembrane segmentand(S1~S12) and the possibility of subcellular localization in plasma membrane was 80%. The deduced polypeptide of SsHAK2 had three transmembrane domains, including potassium transporter domain and amino acid transporter domain. The homology analysis indicated that SsHAK2 shared sequence homology with other members of HAK family in plant, such as Zea mays, Oryza sativa, Hordeum vulgare, with the level of sequence identity ranging from 52% to 95%. The changes in expression of SsHAK2 under low K, drought and salt stresses were detected by qPCR analysis. Under low K stress condition, the gene had the highest inducible expression level at 96 hours, which was 1.70-fold than that of control. Under salt stress, the expression level began to increase rapidly after 48 hours stress of salt and also reached the highest at 96 hours or 4.37-fold than that of control. Under drought stress, the gene had the highest level at 12 hours or 4.07-fold than that of control, and then dropped to 1.69-fold at 24 hours. Afterwards from 24 to 48 hours, it was up-regulated and after 48 hours became down-regulated rapidly, by 96 hours which had the lowest inducible expression level or only 1/4 of that in control. It suggests that SsHAK2 might take part in responses to various stress in addition to low K. This study has established a foundation for future research on understanding the molecular mechanism of K uptake in sugarcane.

Chromosome segment introgression lines (CSILS) are important materials for the study of QTL fine mapping, the interaction between QTLs and mechanism of heterosis. molecular marker assisted selection (MAS) is an important means to identify the introgression segments during the creation of CSILs. In order to clarify the introgression and the lost of alien chromosome segments, and the effects of alien chromosome on the phenotype of Chinese cabbage (Brassica campestris ssp. pekinensis) during the process of establishing Chinese cabbage-non heading Chinese cabbage (Brassica campestris ssp. chinensis) chromosome introgression lines, InDel markers identification and phenotypic analysis of BC2F1 progenies from Chinese cabbage 85-1 and non heading Chinese cabbage JinglvNo.2 were carried out. The results showed that 186 of 264 tested InDel markers from Chinese cabbage were polymorphism between 85-1 and JinglvNo.2, among them, 160 specific markers of JinglvNo.2 compared to 85-1 were screened out, which arranged on 10 chromosomes of B. rapa and the number of InDels on each chromosome varied from 6~26. 52 InDel markers on different position of 10 chromosomes were applied to identify the BC2F1 population. Among them, 50 InDel markers were identified out in 196 BC2F1 plants at different frequency, the plants containing 8 alien chromosome segments were the most common. 40 plants that containing 50 specific markers of JinglvNo.2 and carrying the relatively few alien segments were selected out by analysis of Graphical Geno Typing (GGT) software, which could be further used for the establishment of the introgressed lines with single alien segment. Phenotypic characters of BC2F1 plants, such as leaf shape, leaf size, leaf color and bolting tolerance, flower color showed a serial separation. The results not only provide technical support for the creation of the set of Chinese cabbage-non heading Chinese cabbage chromosome segment introgression lines, but also provide the material for studying the effects of different alien segments on plant phenotypic variation.

Endocrine growth hormone (GH) combining with the growth hormone receptor (GHR) to form GH-GHR-IGF1 pathway, played crucial roles in animal growth. Insulin-like growth factor 1 (IGF1), a kind of regulating polypeptide, acted in animal development and physiological regulation. In this study, the effects of different ways and GH concentration on the expression of IGF1 were analyzed in chicken primary myoblast cells. It was benefit for us to determine the appropriate way and GH concentration to induce IGF1 expression, which laid the basis for further analyzing the regulatory mechanism of GH in muscle development. The chicken (Gallus gallus) primary myoblast cells were isolated and cultured by physical operations and differential adhension methods using chicken embryo hatching to 11 d. The different concentrations of GH (50, 100, 200 and 500 ng/mL) were added into the DuIbecco's modified eagIe's medium (DMEM) media with or without fetal bovine (Bos tarus) serum to culture the cells for 3 h. The cells were harvested and the total RNAs were isolated to analyze the expression of IGF1. Results indicated that chicken GH protein could induce the expression of IGF1 both in mRNA and protein levels. Chicken IGF1 were more highly expressed under the condition of serum starvation than that of serum culture at the same GH concentration. The expression of IGF1 reached the highest level when 200 ng/mL GH was added under the condition of serum starvation. It indicated that the best way to induce IGF1 expression was that adding 200 ng/mL GH into DMEM media after chicken primary myoblast cells were serum starvation for 10 h. The results provide the basic data for broiler muscle development and lays the basis for further studying the roles of GH-GHR-IGF1 pathway in myoblast cells proliferation and differentiation.