Genome-Wide Identification of Tomato (Solanum lycopersicum L.) CKX Gene Family and Expression Analysis in the Callus Tissue under Zeatin Treatment

The cytokinin oxidase/dehydrogenase(CKX)enzyme is essential for controlling thefluctuating levels of endogen-ous cytokinin(CK)and has a significant impact on different aspects of plant growth and development.Nonethe-less,there is limited knowledge about CKX genes in tomato(Solanum lycopersicum L.).Here we performed genome-wide identification and analysis of nine SlCKX family members in tomatoes using bioinformatics tools.The results revealed that nine SlCKX genes were unevenly distributed onfive chromosomes(Chr.1,Chr.4,Chr.8,Chr.10,and Chr.12).The amino acid length,isoelectric points,and molecular weight of the nine SlCKX proteins ranged from 453 to 553,5.77 to 8.59,and 51.661 to 62.494 kD,respectively.Subcellular localization analysis indi-cated that SlCKX2 proteins were located in both the vacuole and cytoplasmic matrix;SlCKX3 and SlCKX5 pro-teins were located in the vacuole;and SlCKX1,4,6,7,8,and 9 proteins were located in the cytoplasmic matrix.Furthermore,we observed differences in the gene structures and phylogenetic relationships of SlCKX proteins among different members.SlCKX1-9 were positioned on two out of the three branches of the CKX phylogenetic tree in the multispecies phylogenetic tree construction,revealing their strong conservation within phylogenetic subgroups.Unique patterns of expression of CKX genes were noticed in callus cultures exposed to varying con-centrations of exogenous ZT,suggesting their roles in specific developmental and physiological functions in the regeneration system.These results may facilitate subsequent functional analysis of SlCKX genes and provide valu-able insights for establishing an efficient regeneration system for tomatoes.

Cloning and Expression Pattern Analysis of Nitrogen- Starvation-induced Genes in Rice

To understand the regulation system of nitrogen X-starvation in higher plants, a cDNA library from N-starved rice (Oryza sativa L.) seedlings was constructed using rapid subtraction hybridization (RaSH) procedure. Through reverse Northern analysis and Northern blotting, 18 unique known genes and two unique unknown genes were identified, which were up-regulated by N-starvation in rice. The known genes are involved in several metabolisms including carbon metabolism, secondary metabolite synthesis, ubiquitylation and protein degradation, phytohormone metabolism, signal transduction, growth regulator and transcription factors. Different induced expression patterns based on spatial and temporal express ions were found for these genes. The results indicate the cross-talks between N-starvation response and various metabolisms in plants.

Temporal and Spatial Expression Patterns of Sox1 Gene in Xenopus laevis Embryo

We describe the temporal and spatial expression pattern of Sox 1 gene during Xenopus laevis early development and compare the expression patterns of Sox 1-3 in the developing eye and brain. Alignment of Sox 1-3 amino acid sequences shows a high conservation within the HMG-box DNA binding domains. RT-PCR analysis indicates that Sox 1 is expressed throughout development from the unfertilized egg to at least the tadpole stage, although at different expression levels. The transcripts of XSox 1 are detected in the animal pole at cleavage and blastrula stages and mainly in the central nervous system （CNS） and the developing eye at neurula stages. The study of the developmental expression of XSox 1 will aid in the elucidation of the function of SoxB 1 subgroup genes in vertebrate neurogenesis.

Assignment and expression patterns of porcine muscle-specific isoform of phosphoglycerate mutase gene

It has been reported that the muscle-specific isoform （type M, PGAM2） of phosphoglycerate mutase （PGAM） is a housekeeping enzyme; it catalyzes the conversion of 3-phosphoglycerate into 2-phosphoglycerate in the glycolysis process to release energy. It is encoded by the Pgam2 gene. In this study, the cDNA of the porcine Pgam2 was cloned. This gene contains an open reading frame of 765 bp encoding a protein of 253 residues, and the predicted protein sequences share high similarity with other mammalians, 96% identity with humans, and 94% identity with mouse and rats. Pgam2 was mapped to SSC18q13-q21 by the RH panel. In this region, there are several QTLs, such as fat ratio, lean percentage, and diameter of muslce fiber, which affect meat production and quality. The reverse transcriptase-polymerase chain reaction revealed that the porcine Pgam2 gene was mainly expressed in the muscle tissue （skeletal muscle and cardiac muscle）, and was expressed highly at skeletal muscle development stages （embryonic periods： 33, 65, and 90 days post-conception （dpo）; postnatal pigs： 4 days and adult）. This indicates that the Pgam2 gene plays an important role in muscle growth and development. In addition, it was demonstrated that PGAM2 locates both in cytoplasm and nuclei, and takes part in the glycometabolism process of cytoplasm and nuclei.

Study on gene expression patterns and functional pathways of peripheral blood monocytes reveals potential molecular mechanism of surgical treatment for periodontitis

BACKGROUND Periodontitis is a chronic inflammation of periodontal supporting tissue caused by local factors. Periodontal surgery can change the gene expression of peripheral blood mononuclear cells. However, little is known about the potential mechanism of surgical treatment for periodontitis. AIM To explore the potential molecular mechanism of surgical treatment for periodontitis. METHODS First, based on the expression profiles of genes related to surgical treatment for periodontitis, a set of expression disorder modules related to surgical treatment for periodontitis were obtained by enrichment analysis. Subsequently, based on crosstalk analysis, we proved that there was a significant crosstalk relationship between module 3 and module 5. Finally, based on predictive analysis of multidimensional regulators, we identified a series of regulatory factors, such as endogenous genes, non-coding RNAs (ncRNAs), and transcription factors, which have potential regulatory effects on periodontitis. RESULTS A total of 337 genes related to surgical treatment for periodontitis were obtained, and 3896 genes related to periodontitis were amplified. Eight expression modules of periodontitis were obtained, involving the aggregation of 2672 gene modules. These modules are mainly involved in G-protein coupled receptor signaling pathway, coupled to cyclic nucleotide second messenger, and adenylate cyclasemodulating G-protein coupled receptor signaling pathway. In addition, eight endogenous genes (including EGF, RPS27A, and GNB3) were screened by network connectivity analysis. Finally, based on this set of potential dysfunction modules, 94 transcription factors (including NFKB1, SP1, and STAT3) and 1198 ncRNAs (including MALAT1, CRNDE, and ANCR) were revealed. These core regulators are thought to be involved in the potential molecular mechanism of periodontitis after surgical treatment. CONCLUSION Based on the results of this study, we can show biologists and pharmacists a new idea to reveal the potential molecular mechanism of surgical treatment for periodontitis, and provide valuable reference for follow-up treatment programs.

Characterization of the imprinting and expression patterns of ZAG2 in maize endosperm and embryo

ZAG2 has been identified as a maternally expressed imprinted gene in maize endosperm.Our study revealed that paternally inherited ZAG2 alleles were imprinted in maize endosperm and embryo at 14 days after pollination(DAP), and consistently imprinted in endosperm at 10, 12, 16, 18, 20, 22, 24, 26, and 28 DAP in reciprocal crosses between B73 and Mo17. ZAG2 alleles were also imprinted in reciprocal crosses between Zheng 58 and Chang7-2 and between Huang C and 178. ZAG2 alleles exhibited differential imprinting in hybrids of 178 × Huang C and B73 × Mo17, while in other hybrids ZAG2 alleles exhibited binary imprinting. The tissue-specific expression pattern of ZAG2 showed that ZAG2 was expressed at a high level in immature ears, suggesting that ZAG2 plays important roles in not only kernel but ear development.

Expression patterns of OCT4, NANOG, and SOX2 in goat preimplantation embryos from in vivo and in vitro

The transcription factors, including OCT4, NANOG, and SOX2, played crucial roles in the maintenance of self-renewal and pluripotency in embryonic stem cells （ESCs）. They expressed in preimplantation mammalian development with spa- tio-temporal pattern and took part in regulation of development. However, their expression and roles in goat had not been reported. In the present study, the expression of OCT4, NANOG, and SOX2 in goat preimplantation embryos both in vivo and in vitro were detected by real-time RCR and immunofluorescence. For in vivo fertilized embryos, the transcripts of OCT4, NANOG, and SOX2 could be detected from oocytes to blastocyst stage, their expression in morula and blastocyst stages was much higher than other stage. OCT4 protein was detected from oocyte to blastocyst, but the fluorescence was more located-intensive with nuclei from 8-cell stage, its expression present in both inner cell mass （ICM） and trophoblast cells （TE） at blastocyse stage. NANOG protein was similar to OCT4, the signaling of fluorescence completely focused on cell nuclei, while the SOX2 firstly showed nuclei location in morula. Comparing to in vivo fertilized embryo, the mRNA of these three transcription factors could be detected at 8-cell stage in parthenogenetic embryos （in vitro）. Thereafter, the expressional level rose gradually along with embryo development. The locations of OCT4 and NANOG proteins were similar to in vivo fertilized embryos, and they located in cell nuclei from morula to blastocyst stage, while SOX2 protein firstly could be detected in cell nuclei at 8-cell stage. These differences suggested that OCT4, NANOG, and SOX2 played different function in regulating development of goat preimplantation embryos. These results may provide a novel insight to goat embryo development and be useful for goat ESCs isolation.

Analysis of Gene Expression Pattern of Lumbar Intervertebral Disc Degeneration in Human

ObjectiveTo investigate the gene expression changes in normal and degeneration lumbar intervertebral disc in humans, providing information for clinical. MethodsThe PCR products of 4096 human genes were spotted onto a kind of chemical-material-coated-glass slides. The total RNAs were isolated from the tissues. Both the mRNAs from the degeneration and normal lumbar intervertebral disc in humans were reversely transcribed to the cDNAs, which used as the hybridization probes with the incorporations of fluorescent dUTP. The mixed probes were then hybridized to the cDNA microarray. After high-stringent washing, the cDNA microarray was scanned for the fluorescent signals and analyzed with computer image analysis. ResultsAmong the 4096 targets, there were 706 genes whose expression levels differed between the degeneration and normal lumbar intervertebral disc in all cases, comprising 298 up-regulated and 358 down-regulated ones. ConclusionDNA microarray technology is an effective technique in screening for differently expressed genes between the degeneration and normal lumbar intervertebral disc. Cell apoptosis plays an important role in the process of lumbar intervertebral disc degeneration.

Identification of SNPs and expression patterns of FZD3 gene and its effect on wool traits in Chinese Merino sheep(Xinjiang Type)

As a member of the Frizzled family, Frizzled3 (FZD3) is a receptor of the canonical Wnt signaling pathway and plays a vital role in mammalian hair follicle developmental processes. However, its effects on wool traits are not clear. The objectives of this study were to identify the single nucleotide polymorphisms (SNPs) and the expression patterns of FZD3 gene, and then to determine whether it affected wool traits of Chinese Merino sheep (Xinjiang Type) or not. PCR-single stranded conformational polymorphism (PCR-SSCP) and sequencing were used to identify mutation loci, and general linear model (GLM) with SAS 9.1 was used for the association analysis between wool traits and SNPs. Quantitative real-time PCR (qRT-PCR) was used to investigate FZD3 gene expression levels. The results showed that six exons of FZD3 gene were amplified and two mutation loci were identified in exon 1 (NC_019459.2: g.101771685 T>C (SNP1)) and exon 3 (NC_019459.2: g.101810848, A>C (SNP2)), respectively. Association analysis showed that SNP1 was significantly associated with mean fiber diameter (MFD)(P=0.04) and live weight (LW)(P=0.0004), SNP2 was significantly associated with greasy fleece weight (GFW)(P=0.04). The expression level of FZD3 gene in skin tissues of the superfine wool (SF) group was significantly lower (P<0.05) than that of the fine wool (F) group. Moreover, it had a higher expression level (P<0.01) in skin tissues than in other tissues of Chinese Merino ewes. While, its expression level had a fluctuant expression in skin tissues at different developmental stages of embryos and born lambs, with the highest expression levels (P<0.01) at the 65th day of embryos. Our study revealed the genetic relationship between FZD3 variants and wool traits and two identified SNPs might serve as potential and valuable genetic markers for sheep breeding and lay a molecular genetic foundation for sheep marker-assisted selection (MAS).

Expression Pattern of Peptide and Amino Acid Genes in Digestive Tract of Transporter Juvenile Turbot(Scophthalmus maximus L.)

Turbot(Scophthalmus maximus L.),a carnivorous fish species with high dietary protein requirement,was chosen to examine the expression pattern of peptide and amino acid transporter genes along its digestive tract which was divided into six segments including stomach,pyloric caeca,rectum,and three equal parts of the remainder of the intestine.The results showed that the expression of two peptide and eleven amino acid transporters genes exhibited distinct patterns.Peptide transporter 1(Pep T1) was rich in proximal intestine while peptide transporter 2(PepT2) was abundant in distal intestine.A number of neutral and cationic amino acid transporters expressed richly in whole intestine including B^0-type amino acid transporter 1(B^0AT1),L-type amino acid transporter 2(LAT2),T-type amino acid transporter 1(TAT1),proton-coupled amino acid transporter 1(PAT1),y^+L-type amino acid transporter 1(y^+LAT1),and cationic amino acid transporter 2(CAT2) while ASC amino acid transporter 2(ASCT2),sodium-coupled neutral amino acid transporter 2(SNAT2),and y^+L-type amino acid transporter 2(y^+LAT2) abundantly expressed in stomach.In addition,system b^(0,+) transporters(rBAT and b^(0,+)AT) existed richly in distal intestine.These findings comprehensively characterized the distribution of solute carrier family proteins,which revealed the relative importance of peptide and amino acid absorption through luminal membrane.Our findings are helpful to understand the mechanism of the utilization of dietary protein in fish with a short digestive tract.

Detection of Cytokine Expression Patterns in the Peripheral Blood of Patients with Acute Leukemia by Antibody Microarray Analysis

The cytokines of acute leukemia （AL） patients have certain expression patterns, forming a complex network involved in diagnosis, progression, and prognosis. We collected the serum of different AL patients before and after complete remission （CR） for detection of cytokines by using an antibody chip. The expression patterns of cytokines were determined by using bioinformatics computational analysis. The results showed that there were significant differences in the cytokine expression patterns between AL patients and normal controls, as well as between acute myeloid leukemia （AML） and acute lymphoblastic leukemia （ALL）. In confirmatory test, ELISA revealed the expression of uPAR in AL. Moreover, the bioinformatic analysis showed that the differentially expressed cytokines among the AL groups were involved in different biological behaviors and were closely related with the development of the disease. It was concluded that the cytokine expression pattern of AL patients is significantly different from that of healthy volunteers. Also, differences of cytokine expression patterns exist between AML and ALL, and between before and after CR in the same subtype of AL, which holds important clinical significance for revealing disease progression.

Genome-wide identification and characterization of the JAZ gene family and its expression patterns under various abiotic stresses in Sorghum bicolor

The jasmonate ZIM domain(JAZ)protein belongs to the TIFY((TIF[F/Y]XG)domain protein)family,which is composed of several plant-specific proteins that play important roles in plant growth,development,and defense responses.However,the mechanism of the sorghum JAZ family in response to abiotic stress remains unclear.In the present study,a total of 17 JAZ genes were identified in sorghum using a Hidden Markov Model search.In addition,real-time quantification polymerase chain reaction(RT-qPCR)was used to analyze the gene expression patterns under abiotic stress.Based on phylogenetic tree analysis,the sorghum JAZ proteins were mainly divided into nine subfamilies.A promoter analysis revealed that the SbJAZ family contains diverse types of promoter cis-acting elements,indicating that JAZ proteins function in multiple pathways upon stress stimulation in plants.According to RT-qPCR,SbJAZ gene expression is tissuespecific.Additionally,under cold,hot,polyethylene glycol,jasmonic acid,abscisic acid,and gibberellin treatments,the expression patterns of SbJAZ genes were distinctly different,indicating that the expression of SbJAZ genes may be coordinated with different stresses.Furthermore,the overexpression of SbJAZ1 in Escherichia coli was found to promote the growth of recombinant cells under abiotic stresses,such as PEG 6000,NaCl,and 40℃ treatments.Altogether,our findings help us to better understand the potential molecular mechanisms of the SbJAZ family in sorghum in response to abiotic stresses.

Expression Pattern and Target Gene of bbu-miR-103-1 in Buffalo(Bubalus bubalis) at Lactation and Non-lactation Periods

[ Objectivel The paper aimed to investigate the expression pattern of bbu-miR-103-1 in buffalo （Bubalus bubalis） at lactation and non-lactation periods, and to predict its target gene and function. [ Method] Expression pattern of bbu-miR-103-1 at lactation and non-lactation periods were detected by qRT-PCR. The precursor expression plasmid of bbu-miR-103-1 was constructed and named LpEZX-pre-miR-103-1. It was packaged and propagated to produce high-titer lenti- virus in 293T cell lines, which could be used to infect buffalo mammary epithelial cells （BMECs） and over express bbu-miR-103-1. The inhibitor of bbu-miR- 103-1 was chemically synthesized and transfected into BMECs to suppress bbu-miR-103-1 at the same time. The relative expression of pantothenate kinase 3 （ PANK3 ） and milk fat metabolism related genes were detected by qRT-PCR. [ Result] The relative expression of bbu-miR-103-1 at lactation period was 5.29 times higher than that at non-lactation period in buffalo （ P 〈 0.01 ）. The LpEZX-pre-miR-103-1 had been successfully constructed and packaged with the infection titer of 3.47×10^6 PFU/mL. Overexpress or suppress of bbu-miR-103-1 extremely down-regulated or up-regulated the expression level of PANK3 in BMECs （ P 〈 0.01 ）. Over expression of bbu-miR-103~l extremely enhanced the expression of Acetyl-CoA carboxylase alpha（ACACA）, Glycerol-3-phosphate acyhransferase 1 mitochon- drial （GPAM）, Diacylglycerol Oacyhransferase l （DGAT1） and Pyrnvate dehydrogenase lipoamide kinase isozyme 4 （PDK4） （P 〈0.01 ）, and also significantly up-regulated the expression of sterol regulatory element binding protein-1 c （SREBPI c）, Adipose differentiation-related protein （ADFP）, Cluster of differentiation 36 （ CD36）, Acetyl-CoA synthetase short-chain subfamily member 1 （ACSS1） （P 〈0.05）. Over expression of bbu-miR-103-1 down-regulated the expression of PANK3, and improved the mRNA level of SREBPlc by feedback regulation, finally promoting the de novo synthesis of fatty acid beginning with ACACA. [ Conclusion] bbu-miR-103-1 plays an important role in enhancing milk fatty acid synthesis, which provides a molecular base for revealing formation and regulatory mechanism of high-level milk fat in buffalo.

Pattern of expression of the CREG gene and CREG protein in the mouse embryo

Background The cellular repressor of ElA-stimulated genes(CREG) is a secreted glycoprotein that inhibits cell proliferation and/or enhances differentiation.CREG is widely expressed in adult tissues such as the brain,heart, lungs,liver,intestines and kidneys in mice.We investigated the level of CREG expression during mouse embryogenesis and its distribution at 18.5 days post coitus(dpc).Methods Immunohistochemical staining with diaminobenzidine,western blotting and reverse transcription-polymerase chain reaction were used.Results CREG expression was rst detected in mouse embryos at 4.5 dpc.It was expressed at almost all stages up to 18.5 dpc.The level of CREG was found to increase gradually and was highest at 18.5 dpc.Western blotting showed that the CREG protein was expressed at higher levels in the brain,heart,intestines and kidneys than in the lungs and liver at 18.5 dpc.In 9.5 dpc embryos,CREG was expressed only in the endothelial cells of blood vessels,after the vascular lumen had formed.With advanced differentiation, vascular smooth muscle cells developed in the embryonic vascular structures;the expression of smooth muscle a-actin protein and CREG were positive and increased gradually in 10.5 dpc embryonic vessels.CREG expression in the embryonic blood vessels peaked at 15.5 dpc and was reduced slightly at 18.5 dpc.Conclusions These results indicate that CREG is expressed during mouse embryogenesis and might participate in the differentiation of these organs during embryogenesis.

Genetic basis of negative heterosis for growth traits in chickens revealed by genome-wide gene expression pattern analysis

Background:Heterosis is an important biological phenomenon that has been extensively utilized in agricultural breeding.However,negative heterosis is also pervasively observed in nature,which can cause unfavorable impacts on production performance.Compared with systematic studies of positive heterosis,the phenomenon of negative heterosis has been largely ignored in genetic studies and breeding programs,and the genetic mechanism of this phenomenon has not been thoroughly elucidated to date.Here,we used chickens,the most common agricultural animals worldwide,to determine the genetic and molecular mechanisms of negative heterosis.Results:We performed reciprocal crossing experiments with two distinct chicken lines and found that the body weight presented widely negative heterosis in the early growth of chickens.Negative heterosis of carcass traits was more common than positive heterosis,especially breast muscle mass,which was over−40%in reciprocal progenies.Genome-wide gene expression pattern analyses of breast muscle tissues revealed that nonadditivity,including dominance and overdominace,was the major gene inheritance pattern.Nonadditive genes,including a substantial number of genes encoding ATPase and NADH dehydrogenase,accounted for more than 68%of differentially expressed genes in reciprocal crosses(4257 of 5587 and 3617 of 5243,respectively).Moreover,nonadditive genes were significantly associated with the biological process of oxidative phosphorylation,which is the major metabolic pathway for energy release and animal growth and development.The detection of ATP content and ATPase activity for purebred and crossbred progenies further confirmed that chickens with lower muscle yield had lower ATP concentrations but higher hydrolysis activity,which supported the important role of oxidative phosphorylation in negative heterosis for growth traits in chickens.Conclusions:These findings revealed that nonadditive genes and their related oxidative phosphorylation were the major genetic and molecular factors in the negative heterosis of growth in chickens,which would be beneficial to future breeding strategies.

Bioinformatics and Expression Pattern Analysis of Tomato ns LTP 2-like cDNA full-length Gene Clone

TDF1(transcription-drived fragment) was homologous to the predicted S. lycopersicum nonspecific lipid-transfer protein,nsLTP 2-like(91%), and it was significantly upregulated in response to C. fulvum(cladosporium fulvum) infection in tomato plants.In this experiment, the full-length cDNA of nsLTP 2-like was cloned using RACE technology based on the sequence of TDF1(GenBank: JZ717725). A full-length, 625 bp(GenBank: KU366289), cDNA sequence, which with 98% similarity to nsLTP 2-like gene(GenBank: XM015233692) was obtained. This cDNA contains an ORF(open reading frame) with full-length of 345 bp, coding of 114 amino acids, including 12.3% Ala and Gly. Protein molecular weight was 11.51 ku, the isoelectric point(pI) was 8.99, and average overall hydrophilicity was 0.412, with one phosphorylation sites, belonging to volatile acidic nuclear protein. Secondary structure prediction showed that α-Helix accounts for 30.7%, extension chain for 12.28%, β-corner for 9.65%, and random coil for 47.37%. Through comparative analysis of the homology among species, it was found that the amino acid sequence of tomato nsLTP 2-like protein had a high similarity with other plants, and with a specific conserved sequence which might related features in nsLTP 2-like protein. It also be analyzed the gene expression pattern of tomato in different parts and under different stress conditions.The results showed that nsLTP 2-like gene was up-regulated in varying degrees, under the condition of cold stress, exogenous hormone spraying and cladosporium fulvum infection. Therefore, it was speculated that the gene played a role in response to abiotic and biotic stress in tomato.

Molecular Characterization and Expression Pattern of Rheb Gene in Inner Mongolia Cashmere Goat(Capra hircus)

As one member of the Ras super family, Rheb is an upstream regulator of mTOR signaling pathway, which regulates the process of cell-growth, proliferation and differentiation. In order to study the relationship between Rheb and mTOR in Inner Mongolian Cashmere goat （Capra hircus） cells, Ras homolog enriched in brain （Rheb） gene eDNA was amplified by RT-PCR. It is 555 bp in length and includes the complete ORF encoding 184 amino acids （GenBank accession no. HM569224）. The full eDNA nucleotide sequence has a 99% identity with that of sheep, 98% with cattle and 93% with human while their amino acids sequence shares identity with 98, 97 and 97% of them, correspondingly. The bioinformatics analysis showed that Rheb has a Ras family domain, two casein kinase II phosphorylation sites, two ATP/GTP-binding sites motifA （P-loop）, a prenyl group binding site （CAAX box）. Tissue-specific expression analysis performed by semi- quantitative RT-PCR. The Rheb gene was expressed in all the tested tissues and the highest level ofmRNA accumulation was detected in brain, suggesting that Rheb played an important role in goat cells.

Cloning, Expression Pattern Analysis and Subcellular Localization of Resveratrol Synthase Gene in Peanut (<i>Arachis hypogaea</i>L.)

Resveratrol synthase (RS) is a key enzyme that plays a critical role in the resveratrol synthesis pathway. In this study, six RS genes were isolated and characterized from peanut variety “Zhenzhu Hong” by silico cloning and RT-PCR. Bioinformatics analysis showed that deduced amino acid sequences of the six cloned RS genes were highly conserved with a similarity from 95% to 99% when compared to the RS genes which had been deposited at the GenBank. The results of amino acid sequences analysis showed six RS proteins contained the Chal_Sti_Synt_N and ACP_Syn_III_C domains and can be classified to same family but with different evolutionary distance. Expression pattern analysis by QRT-PCR provided evidence indicating that the mRNA of six RS genes were primarily expressed in the peanut shell at different developmental stages with different expression levels, but only lower levels of them were evident in the peanut kernel. The subcellular localization of RS protein in onion epidermal cell was performed by Agrobacterium tumefaciens-mediated transformation and the green fluorescent was monitored by confocal fluorescence microscopy. The results indicated that, RS1 and RS5 were located in the nucleus and plasma membrane respectively, while the RS2, RS3, RS4 and RS6 were located in both nucleus inner membrane and plasma membrane. The data will provide basic information for elucidating the regulatory mechanisms and enzyme kinetics underlying the RS genes in the resveratrol synthase pathway.

Molecular Cloning, Expression Pattern, and 3D Structural Prediction of the Cold Inducible RNA- Binding Protein(CIRP) in Japanese Flounder(Paralichthys olivaceus)

Cold-inducible RNA-binding protein(CIRP) is a kind of RNA binding proteins that plays important roles in many physiological processes. The CIRP has been widely studied in mammals and amphibians since it was first cloned from mammals. On the contrary, there are little reports in teleosts. In this study, the Po CIRP gene of the Japanese flounder was cloned and sequenced. The genomic sequence consists of seven exons and six introns. The putative Po CIRP protein of flounder was 198 amino acid residues long containing the RNA recognition motif(RRM). Phylogenetic analysis showed that the flounder Po CIRP is highly conserved with other teleost CIRPs. The 5' flanking sequence was cloned by genome walking and many transcription factor binding sites were identified. There is a Cp Gs region located in promoter and exon I region and the methylation state is low. Quantitative real-time PCR analysis uncovered that Po CIRP gene was widely expressed in adult tissues with the highest expression level in the ovary. The m RNA of the Po CIRP was maternally deposited and the expression level of the gene was regulated up during the gastrula and neurula stages. In order to gain the information how the protein interacts with m RNA, we performed the modeling of the 3D structure of the flounder Po CIRP. The results showed a cleft existing the surface of the molecular. Taken together, the results indicate that the CIRP is a multifunctional molecular in teleosts and the findings about the structure provide valuable information for understanding the basis of this protein's function.

Preliminary analysis of two NAC transcription factor expression patterns in Larix olgensis

The NAC transcription factor family is plant-specific with various biological functions.However,there are few studies on the NAC gene involving coniferous species.Bioinformatics research and expression analysis of NAC genes in Larix olgensis can be used to analyse the function of the NAC gene in the future.Screening of excellent genetic materials and molecular breeding have been utilized to cultivate high-quality,stress-resistant larches.According to the transcriptome data for L.olgensis,the genes Uni-gene81490 and Unigene70699 with complete ORFs(open reading frames)were obtained by conserved domain analy-sis and named LoNAC1 and LoNAC2,respectively.The cDNAs of LoNAC1 and LoNAC2 were 1971 bp and 1095 bp in length,encoding 656 and 364 amino acids,respectively.The molecular weights of the proteins encoded by the two genes were predicted to be 72.61 kDa and 41.13 kDa,and subcellular localization analysis indicated that the proteins were concentrated in the nucleus.The results of real-time quantitative PCR analysis showed that at different growth stages and in different tissues of L.olgensis,the relative expression levels of the two NAC genes were highest in the stem,and the expression differences were more obvious in non-lignified tissues.After drought,salt and alkali stress and hormone treatment,expression was induced to different degrees.The expression levels of LoNAC1 and LoNAC2 in semi-lignified L.olgensis were higher than in the other two periods(non-lignified and lignified),and expression levels significantly increased under drought and salt stress.Relative expression levels changed under hormone treatment.It is speculated that these two genes may not only be related to drought and salt stress and secondary growth but may also be induced by hormones such as abscisic acid.Overall,LoNAC1 and LoNAC2 are genetic materials that can be used for molecular breeding of larch.