The genetic diversity of the diazotrophic population in the rhizosphere soil and the washed roots of rice was investigated using nifH PCR—RFLP. The PCR—RFLP profiles were compared and analyzed with UPGMA and by maximum parsimony clustering. The RFLP profiles with the restriction enzyme Mnl showed that there are eight the RFLP pairs of similar patterns (similarity>50%) and two pares of similar patterns with 100% similarity between the rhizosphere soil and the root surface/tissue; and three specific patterns generated from the rhizoshere soil, and the washed root respectively, implying high diversity of the diazotrophe presents in the rice rhizoshpere soil and the washed rice roots of rice.

The rice blast fungus Magnaporthe grisea causes one of the most destructive diseases of rice around the world. Mutant generation is the basic work to study mutation mechanism of Magnaporthe grisea. Based on the transformation system established by our laboratory, A large scale of mutants were generated through T-DNA insertion Agrobacterium tumifaciens-mediated transformation(ATMT), in which some morphology and pathogenicity mutants were obtained. In the paper, we selected 28 mutants from among those mutants for further research, including morphology and development observation. Within 28 selected mutants, it was found that there were 15 color abnormal in colony mutants, 8 decreased growth rate mutants, 2 abnormal in conidium morphology and sporulation mutants, 2 abnormal appressorium mutants and 3 appressorium defective mutants. By inoculation the conidia of the mutants on the susceptible rice variety Nipponbare and resistant variety C101(as control). Of the 28 mutants tested, 9 showed completely pathogenicity lost and disease grade 0. By cross cultivation of the mutants with the mating type testers (P131 and 1528), we found 3 mutants(Y34-0211,Y34-1469 and Y34-0635) completely lost the ability to turn their anamorph into teleomorph phase, while this ability of other mutants tested remained the same as the wild type.

For mapping and characterizing QTLs related to Phytophthora capsici (PC) resistance in pepper, a molecular linkage map was constructed with AFLP markers in an intraspecific F2 generation population by crossing 93-100-17-1-0, a highly resistant to PC, with Qiemen, an inbred line susceptible to PC. A total of 165 F2 generation plants from the cross between 93-100-17-1-0 and Qiemen were used for identification of DNA markers linked to PC resistance. The results of the genetic statistical analysis using three different criteria, receptivity, inductivity, and stability, indicated that the resistance of 93-100-17-1-0 to PC was controlled by polygenes or oligogenes. A genetic map was constructed using the software Mapmaker/exp, which contained 11 linkage groups with 46 linked molecular markers and covered 1381.5 cM of the whole genome. Using the three different criteria as traits, 18 QTLs localizing on LG1, LG2, LG5, LG7 and LG8, which explained over 64% of the phenotypic variance, were identified using the Function Interval mapping and Composite Interval mapping of the software winQTLCART. The most significant QTL localized on LG5 from 128 cM to 132 cM, which explained 68% of the phenotypic variance and whose genetic distance was only 4 cM.

The genetic diversity and genetic variation within and among four tea populations in Hunan were studied by RAPD markers. The result showed that 226 loci were detected by using 21 random primers (10bp) of which 162 loci were polymorphic (88.9%). Species genetic diversity indicated by Shannon's index was 0.43, 74.7% of which was due to within population genetic diversity and 25.3% among population variation. Species gene diversity indicated by Nei's index was 0.37, gene diversity within populations (HS) was 0.28, gene diversity among populations (HST) was 0.09 and the coefficient of gene differentiation (GST) among the populations was 0.23, 76.7% of which was within populations and 23.3% was among populations. The gene diversity estimated by Nei's index was consistent with that estimated by Shannon's index. These results suggest that there is rich genetic within population variation, which offers excellent prospects for seeding selection. The estimate of gene flow from GST (Nm) was 0.74, which indicated quite small genetic recombination between populations, it probably is related to environmental adaptations of four tea populations in Hunan and population isolation in their high mountain habitat.

DSN (duplex-specific nuclease ) displays a strong preference for cleaving ds DNA and DNA in DNA–RNA hybrid duplexes compared with ss DNA and RNA irrespective of sequence length , as described recently. The technique termed DSN is a simple and effective cDNA normalization method. In order to screem and isolate new and full-length sporulation-specific genes of entomopathogenic fungus Metarhizium in the sporulation stage with great efficiency, a combined DSN normalization method with the SMARTTM (switching mechanism at 5′end of RNA transcript) library construction method was used to construct a normalization and full-length cDNA library of entomopathogenic fungus Metarhizium in the sporulation stage. The titer of unamplified cDNA library was 2.1 ×106 cfu/mL and contained 6×106 independent clones.The average cDNA inserts size was 1500bp with a recombination rate of 95%.Sequencing results and bioinformatics analysis of random picked clones indicated that the ratio of full-length cDNA was about 60％.The presentation levels of the abundant transcripts G3PDH and β-tubulin decreased obviously compared with non-normalized samples.

This experiment was conducted to study the effect bovine lactoferrin (bLF) on antimicrobial peptide PMAP-37 gene expression. In vivo, seventy-two Duroc × Landrace × TaiHu weaning piglets (♂ Duroc× ♀Landrace× TaiHu) of 36 days were divided into three treatment groups randomly by weight and each group included three repeats of eight piglets. The control (treatment 1) received the basal dietary without bLF. Treatment 2, treatment 3 received dietary contained 0.125% bLF and 0.25% bLF respectively. In vitro, we separated bone marrow cells and cultured with bLF with 10, 100, and 1000 µg/ml respectively for 3h and 6h, and each treatment repeated three times. The control wells were incubated with culture medium. The primers for PMAP-37 and 18S rRNA were designed respectively according to the reported porcine antimicrobial peptide PMAP-37 gene and housekeeping gene 18S rRNA sequences, the suitable MgCl2 concentration and cycles in PCR system were discussed. So an optimized semi-quantitative RT-PCR was constructed, 18S rRNA used as inner control, to detect effect of different concentration bLF on antimicrobial peptide PMAP-37 gene expression in diet and in bone marrow cells of pigs. The results indicated that, compared with control group, 0.125% bLF and 0.25% bLF in diet increased the gene expression of antimicrobial peptide PMAP-37 by 109.62% (P<0.01) and 69.23% (P<0.05) respectively. In bone marrow cells, 10, 100, and 1000 µg/ml bLF also increased gene expression of PMAP-37 respectively for 3h and 6h, but statistic result was not significant. These observations suggest that antimicrobial peptide PMAP-37 gene expression could be regulated by bLF.

To identify the aluminum stress induced genes of alfalfa, a forward subtractive cDNA library was constructed from AL-treated and non-treated seedling roots by suppression subtractive hybridization. Colony PCR suggested that the iniserts were between 250bp~1200bp and the library was suitable for the following work. 40 clones were randomly selecteed, secquenced and obtained 36 unique ESTs. BLASTX show that these ESTs including antioxidative stress, signal transduction, development and energy metaboly, liake peroxidase, calcium-dependent protein kinase, neutral/alkaline invertas, protein phosphatase regulator ect. Many ESTs related to other stresses were also found in our study and nine ESTs had no homology from NCBI.

Abstract: To establish an ISSR marker targeted for D. villosum specific chromosome, the PCR analysis was performed on D. villosum, Chinese Spring, Mianyang11, Triticum tetres and other materials by 96 ISSR (inter-simple sequence repeat) primers for screening. A specific segment of 200~500bp was amplified

The retinoid (vitamin A and its derivatives) plays a critical role in vision, growth, reproduction, cell differentiation, and embryonic development. Using the INRA IMpRH panel, porcine cellular retinol binding protein genes 1 (RBP1) and plasma retinol binding protein gene 4 (RBP4) were assigned to porcine chromosomes 13 and 14, respectively. The mRNA distributions of the two genes in adult Wuzhishan pig tissues (lung, skeletal muscle, spleen, heart, stomach, large intestine, lymph code, small intestine, liver, brain, kidney, and fat) were examined. One SNP was identified in PRB4, and analyzed using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method in Laiwu, Wuzhishan, Guizhou, Bama pig breeds and Tongcheng experimental population. Significant associations were found between different genotypes of RBP4-A/G156 with maximum carcass length (LM), minimum carcass length (LN), hemoglobin (HGB), mean corpuscular volume hemoglobin (MCH), and day to slaughter weight (Age) traits. This SNP locus may be used as the genetic markers in pig breeding and porcine production.

The Zygote arrest 1(ZAR1) gene （gi:83727928） was employed to investigate variation character and associate with reproductive traits in five pig purebreds (Small Meishan, Qingping, Duroc, Landrace and Large White). We identified six new SNPs in the second and third exons, addition with one 5 bp deletion/insertion mutation in the third intron in the ZAR1 gene by using PCR-SSCP and DNA sequencing technique. Exon3-BstU I (C54/T54) located in the 54th of exon3 and Intron3-TspR I (5 bp deletion/insertion mutation) in intron3 were genotyped and associated with reproductive traits in these five pig breeds. Association results showed that Exon3-BstU I polymorphism had a significant effective （P<0.05）with total number born in first litter TNB1)and total number born in second litter(TNB2). Total number born (NBA) with CC genotype had more piglets than that of other genotypes in the Exon3-BstU I in the Duroc. Intron3-TspR I polymorphism had a significant effective （P<0.05）with total number born in first litter (TNB1)and total number born in later litters(TNB2) in the Duroc.

The target sequence in RSTN-T plasmid that contained pig resistin cDNA was amplified by PCR,inserted into eukaryotic expression vector pcDNA3.1(+), and then identified by restriction mapping and sequencing.The recombinant expression vector was transformed into Hela cell by LipofectaminTM 2000. The G418-resistant Hela cell clones were selected as well as the RSTN mRNA and the protein expression were analyzed by RT-PCR and Tricine-SDS-PAGE、Western blot,respectively. DNA sequencing demonstrated that pcDNA3.1-RSTN recombinant vector was constructed successfully. Hela cell clones were got by G418 selecting.The expression of pcDNA3.1-RSTN at mRNA level and protein level inHela cell was confirmed by RT-PCR and Tricine-SDS-PAGE、Western blot respectively. The successful construction and expression of pcDNA3.1-RSTN provides foundation for the further study of the pig resistin DNA vaccines.

Abstract: Purified PM-VP2-E290 and Bac-N-Blue DNA were cotransinfected insect cell sf9. The recombinant baculovirus were selected by their lacZ phenotypes and plaque-purified until no more wild type virus could be detected. Then, high-titer viral stocks of the recombinant baculovirus were prepared and used for expression. After SDS-PAGE analysis, an extra band of 67KDa was observed in the extracts of infected sf9 insect cells.The chimeric proteins had strong positive reactions with PPV specific antibody and E290-specific antibody. A large number of virus-like particles formed by expressed protein were observed under Immuno-electron microscopy (IEM).The BALB/c mice injected with the expressed protein, in the absence of adjuvant. showed that the hybrid recombinant parvovirus like-particles formed by the self-assembly of the VP2 capsid protein of PPV carring the CSFV epitope at its N terminal, not only induced a strong CD8+ class restricted CTL response, But stimulate a highly PPV specific antibody levels. Moreover, the antibody levels were significantly higher than that of the inactivated vaccine control.

A real time RT-PCR detection of Bovine central nervous system tissue （CNS）as bovine spongiform encephalopathy（BSE）specified risk material （SRM） in beef and beef products based on glial fibrillary acidic protein (GFAP) mRNA was reported in this study. The results showed that the developed method allowed the detection of CNS tissues from bovine and ovine origins, but not from porcine and avian origins, and that the lowest detection limit was below 0.001% bovine brain homogenate, and the GFAP mRNA signal detection was not affected by 100°C heating treatment for 30min and long period of storage at over 30°C room temperature (RT) for 4 days and at 4°C for 15 days. It is concluded that real time RT-PCR based on GFAP mRNA can serve as a sensitive and specific test in routine inspection and quarantine detection for illegal use of bovine CNS tissues in beef and beef products.

Abstract: To understand the genetic basis of seed setting (SS) in rice, genotype by environment (G×E) interaction and QTLs for SS were analyzed using a population with 202 recombinant inbred lines (RIL) derived from a cross between 2 indica varieties, T226 and T219, under eight different environments. Significant G×E interaction for SS in rice was detected using AMMI statistical model, and the IPCA1 and IPCA2 of G×E interaction explained a total of 57.6% of the variation. QTLs controlling the SS were detected by interval analysis, and a total of 17 QTLs on 9 chromosomes were identified across eight environments, explained the phenotypic variances from 4.6% to 35.7%, respectively. Most of the QTLs were detected in one or two environment, explained small part of phenotypic variances. While a QTL（qSS3-1） on chromosome 3 detected across six environments, explained maximum phenotypic variances in each detected environment and its allele for increasing the SS was derived from T226. The QTL (qSS5-3) on chromosome 5 was detected in 5 environments, however Its allele from T219 increasing the SS .

Abstract: One novel vegetative insecticidal protein (vip) gene, vip3A-LS1, was cloned and identified as GenBank accession number DQ016968 (Vip3Aa22). Alignment results showed that the Vip3A-LS1 deduced protein had 8 amino acids that were different from published homologous proteins. Vip3A-LS1 inserted into pQE30 and transformed into Escherichia coli M15 was expressed to a high level after being induced by IPTG. The intracellular soluble protein extracted from the transformant possessed high toxicity to the larvae of Helicoverpa armigera and Spodoptera exigua, and the LC50 of its purified protein was 73.6 ug/g to H. armigera, and 32.2 ug/g to S. exigua respectively.

BALB/c mice were immunized intraperitoneally with purified recombinant pET-mEIL-18 protein. Murine myeloma cells were fused with the splenocytes of the immunized mice after the third immunization. An indirect ELISA with recombinant mEIL-18 come from baculovirus expression system as antigen was used to screen antibody-producing hybridomas. 12 McAbs against pET-mEIL-18 were obtained. Nine hybridomas cells of them can produce McAbs steadily after 3 cycles of cloning and all of them showed positive reaction to mEIL-18 in IFA tests. The 9 McAbs were reactive with pET-mEIL-18-(1) or pET-mEIL-18-(2) respectively in specific ELISA tests. As a result, five McAbs of them showed positive reaction to pET-mEIL-18-(1) protein, others show positive reaction to pET-mEIL-18-(2) protein. Three McAbs are IgM isotype and the others belong to IgG1 isotype, the light chain of all McAbs is 僴chain. The McAbs against pET-mEIL-18 protein developed in the study would be useful as a basis of detection methods for mEIL-18, investigating the structure and biological effects of mEIL-18.

Single nucleotide polymorphisms (SNPs) of hircine GHRH and INHBA gene loci were analyzed by PCR-SSCP in Boer goat and Xuhuai white goat, and detected the polymorphism distributions of these two loci. Heterozygosity (H), effective number of alleles (Ne), polymorphism information content (PIC) and fixation index (F) were calculated. All loci obey Hardy-Weinberg equilibrium.

Follistatin inhibits the biological action of several TGF-β family members and promotes muscle growth. In this research largemouth bass Follistatin cDNA was isolated. First, total RNA was isolated from ovary of adult female largemouth bass, and then used to clone the Follistatin cDNA by using RT-PCR and rapid cDNA end amplification. The sequence analysis results showed that the full-length of largemouth bass Follistatin was 1444 bp with an open reading frame(ORF) of 966 bp. The deduced ORF amino acid sequence contained a putative signal peptide of 31 amino acids and a mature peptide of 290 amino acids that was composed of four domains including N-domain, DomainⅠ, Domain Ⅱand Domain Ⅲ. Comparing Follistatin mature peptide of largemouth bass with those of torafugu, grass carp, zebrafish, Atlantic salmon and channel catfish, the results showed that the amino acid homology is 97％, 89％, 88％, 88％ and 70％, respectively. Follistatin protein binds with several TGF-β family members through its N-domain. Comparing Follistatin N-domain of largemouth bass with those of torafugu, grass carp, zebrafish, Atlantic salmon and channel catfish, African clawed toad, human, pig, rat and chicken, the results showed that the amino acid homology is 75%~100％. It was clear that the sequences of N-domain showed more conservation than those of mature peptide, which suggested that it was highly restricted in the process of evolution and its functional importance. In order to gain Follistatin fusion protein, Follistatin mature peptide cDNA was inserted into pET-32a(+) vector, which was transformed into BL21 and induced by IPTG. Expression products were detected by SDS-PAGE and confirmed by Western blotting analysis, which showed that the molecule weight of Follistatin fusion protein was about 52kD. This study will benefit further researching of Follistatin promoting muscle growth.

The mitochondrial functional protein alternative oxidase （AOX）genes were took as the research target in this experiment, the genes of all strains that preserved in our lab were amplfied and sequenced. Then compared the sequences with other related sequences downloaded from GenBank by using Clustal X1.81.Three types of phylogenetic analysis were used on the aligned sequences to assess relationships among isolates, the distance-based neighbor-joining(NJ) analysis, parsimony(MP) analysis and Maximum Likelihood(ML). NJ tree and MP trees were constructed with the program PAUP, while ML tree was made by using PUZZLE Version 4.1.Then determined phylogentic realationship of different Cryptospoiridum genus or genotype and contrasted those phylogentic trees with that of drawed by 18SrRNA and HSP70 gene sequence, and determined whether the AOX gene is suitable for phylogentic analysis of Cryptosporidim or not. In those phylogentic trees Cryptosporidium species formed two group, one group contained C.baileyi and C.meleagridis , the other group contained C.hominis、C.suis、C.parvum cattle genotype and C.parvum mouse genotype. The sequence identity of different Cryptosporidium species ranges from 91.8% to 99.6%. The results showed that the AOX gene is equally suitable for the phylogentic analysis of Cryptosporidim .

Anthers from the Fl hybrids of eggplant（Solanum melongena L.）were used as the experimental material. The calli, embryoids and plantlets were obtained through isolated microspore culture. The results were as follows: (1) Most of calli and embryoids were derived from division of vegetative cell of cultured microspores but a part of calli and embryoids were developed from the equal division of microspores, The reproductive cell divided only 1-2 times .(2) The multicellular masses were released from the pollen ditch or the germinal aperture of microspores. (3) The anther tissue was good for embryogenesis in isolated microspores. (4)The chromsome number was examined in cells of root tips of 37 randomly selected regenerations among which 22 were haploids, 14 diploids and 1 tetraploids.

A bioassay method was developed to use parasporal crystal protein of Bacillus thringiensis against plant-parasitic nematodes. Using this method, the parasporal crystal proteins of 10 Bt strains were determined to have activity against plant-parasitic nematodes. The toxicity of YBT-021 against Meloidogyne hapla, Pratylenchus scribneri, Tylenchornchus sp. and Ditylenchus destructor were also assayed. LC50 values were 35.62μg/mL， 75.65μg/mL， 94.31μg/mL and 215.21μg/mL, respectively.

Effects of different DHEA supplemental levels in Arbor Acres broiler feed on lipid metabolism and related hormones were studied in this paper. 180 day-old Arbor Acres broilers were randomly allocated to three groups, each group included three replicates with 20 broilers each. DHEA was not added in the basal feed (control group). The low DHEA and high DHEA treated groups were fed with the basal feed with the supplementation of 5 and 20 mg•kg-1DHEA from 1 to 42 day-old, respectively. Twelve cocks and hens per group were randomly chosen for blood, liver, Abdominal fat, left breast and thigh muscle samples at 42 day-old. The content of blood glucose(BG), triglycerides (TG), total cholesterol (TC), high density lipoprotein- cholesterol (HDL-C), low density lipoprotein-cholesterol (LDL-C),non-esterified fatty acid (NEFA) ,T3,T4,FT3,FT4, glucagon , leptin and the activity of LPL were determined. The result showed that the body weight , abdominal fat weight, percentage of abdominal fat weight and triglycerides(TG), total serum cholesterol (TC), high density lipoprotein-cholesterol (HDL-C), FT3,FT4 were significantly decreased (P＜0.05)and leptin was significantly increased by DHEA in both gender . the content of blood glucose(BG) and LDL-C were not affected by DHEA. The content of NEFA was obviously elevated in the cocks and decreased in the hens (P＜0.05) by DHEA. The activity of LPL and the content of glucagon in serum were significantly enhanced and T4 was significantly decreased in the cocks(P＜0.05)by DHEA. The results implied that DHEA regulated lipid metabolism of broilers by enhancing the enzyme activity and hormone levels related to lipolysis.

The effects of ultraviolet radiation ( 0-150 mj﹒cm-2) on the growth, development and reproduction of Moina mongolica Daday were studied. Two experiments were conducted in laboratory. The results showed that M. mongolica had been injured seriously and its fecundity were decreased in high dosage of UV radiation (above 70 mj cm-2) e.g. pre-reproduction period , neonates number per brood and fecundity was 5.8d, 13 ind. brood-1 and 186 ind. respectively. And relate indicates were 5.4-5.6d, 23.9-33.1ind. brood-1 and 406.8-535.8 ind. respectively, in other treatments groups ( UV 60, 50 ,40 ,30 ,20,10 mj cm-2) and control group. The capability of growth or reproduction of M. mongolica , such as life fecundity and daily increase rate of body length could be stimulated by low dosage of UV radiation. E.g. above two indicates in treatment group were higher than those of control grops in experiments 1; the cumulative duration of development of adult instar of M. mongolica in treatment groups was less than that of control groups in experiment 2. The effects of UV radiation on the life of aquatic organisms and use prospect in mass culture of M. mongolica were discussed.

Suppression subtractive hybridization was used to compare gene expression of middle versus late lactation of Xinong Saanen Goat mammary gland, and the goat butyrophilin(gBTN1A1) gene was screened form subtracted cDNA library of difference expressed genes. After electrical spilicing, The gBTN1A1 gene was amplified and cloned from total RNA of goat mammary gland by RT-PCR , and Registered in Genbank. Sequence analysis indicated that the gBTN1A1 complementary DNA that contained an open reading frame of 1581 nucleotides encoding a putative protein of 526 amino acids was identified. The whole DNA sequence includes 7 exons and 6 introns and amino acids through 1-26 constitute the signal peptide domain. The homologies of nucleotide and peptide sequence of the butyrophilin with bovine (NM_174508), human (NM_001732) and mouse’s (AK145168) are 97%, 88%, 84%，96%, 84% and 70%, respectively. The predicted structure of gBTN1A1 was similar to that of bovine and human, although some differences exist in transmembrane domain and hydrophilicity among them. Our results suggested that the goat mammary gland mRNA level of gBTN1A1 was not only responsible for milk fat globule secretion, but also for milk yield.

PCR primers were designed based on the conserved domain of Arabidopsis WRKY transcription factor, the full-length cDNA of WRKY transcription factor was cloned from Brasssica campestris ssp.chinensis using RT-PCR and RACE techniques. Sequence analysis indicated that BcWRKY1 gene(GenBank accession number: AY836002) consisted of 980 nucleotides(nt), and deduced amino acid sequence containing 285 amino acids. Further comparison showed its identity was 74% and 59% to AtWRKY18 gene and AtWRKY60 gene of Arabidopsis, respectively. The deduced amino acid sequence of WRKY1 gene has lower homology with other plant WRKY genes. Southern-blot analysis indicated there was a single copy in genome of B. campestris ssp.chinensis. Semi-quantitative RT-PCR demonstrated that the corresponding mRNA of WRKY1 was accumulated most abundantly 2h-8h after treatment upon 2mmol/L SA, and 6h-12h after infection by Peronospora parasitica, respectively.

Alpha-Dioxygenase 2 is obtained from ABRC,the coding sequence is amplified using primers containing SmaI and XhoI digest cites,then inserted into pMD-T simple vector,make sure it is right by sequencing,DOX2 is inserted into pGEX-5X-1 by double digest and ligation,then it is transformed into BL21 （DE3） pLysS and BL21 (DE3)-RIPL codon +,DOX2 is expressed efficiently in BL21 (DE3)-RIPL codon + by SDS-PAGE and Western blot analysis,further solubility analysis indicates most of the recombinant protein is expressed in the form of inclusion body,soluble part is harvested by GST affinity purification,and it is used to activity test.Bioinformatics analyisis indicates: DOX2 gene has 88 rare codons in E.coli,nearly 14% of the total codons.Subcellular prediction by SVM indicates:the enzyme is likely to locate in cytoplast.

It is important to understand the mechanism of wheat tolerance to drought, to investigate the wheat transcriptional profiling drought stress, a drought tolerance variety-Hanxuan 10 was treated by PEG6000,RNA samples were collected at 0, 1, 6, and 24 hours respectively, influence dye-labeled complimentary DNA were used to hybridize with the whole genome rice gene chip, which contains over 60K oligos designed based on rice sequence. Data analysis indicated that the number of differentially expressed genes was increased as longer PEG treatment, functional category analysis results indicate that number of energy metabolism pathway-related gene was increased and account for more portion of differentially expressed genes. Most of the photosynthesis related genes was up-regulated but interestingly, Psbr and Rubisco coding genes were down-regulated, suggesting their potential role in drought tolerance response.

Abstract: Using a RIL population derived from a cross between a common wheat 3338 and spelt wheat Altgold, a genetic linkage map was constructed comprising of 287 out of totally 344 polymorphic markers （271 SSR and 63 EST-SSR markers）. Among the 334 polymorphic markers, 82 markers showed the significantly segregation distortion (P＜0.05), favoring either the maker alleles of female parent 3338(33, 40.2%) or male parent Altgold (49,59.8％). Segregation distortion markers distribution along the present molecular maps of wheat was far from uniform, with clusters of tightly linked loci and single markers. 6 segregation distortion regions (clusters of 3-6 makers) were detected on 5 chromosomes, indicating that possible causes for segregation deviation of molecular markers are gametic selection.

【OBJECTIVE】 to establish model of piglet testis tissue in vitro spermatogenesis;【METHOD】Using 5＇-Bromo-2＇- deoxyuridine（BrdU）immunofluorescence technique to detect the proliferation and differentiation of germ cells from testis tissue..And,in this research the Hematoxylin-eosin staining(HE) was also used to identify the differentiation degree ；【RESULTS】As a result,we found that on Day 3 of in vitro culture,sertoli cells began to adhere to the dish;on Day 4~8 of in vitro culture,the germ cells began to travel from the tissue, yet no proliferation or differentiation can be seen；on Day 9~20 of vitro culture , proliferation or differentiation can be clearly observed ,germ cells were linked by the gap junction presented as grape cluster or the strings of beads, and the most exiting thing was that we also found the spermatid in the dish;【CONCLUSION】germ cells can polify or differentiate in this system,and the most interesting thing is that germ cells can differentiate into spermatids without addition of testosterone.

Lipoprotein lipase（LPL）is a key enzyme for the metabolize of circulating triacylglycerols. It is considered to be the major candidate gene affecting fat thickness and intramuscular fat content etc. In this study, PCR-RFLP was applied to analyze the polymorphism of LPL gene in 238 cattle that comprised of six bovine breeds, namely Angus、Hereford、Chinese simmental、Charolais、 Luxi cattle、simmental×Mongolia cattle. A 378 bp-long PCR product was digested with MspⅠ. The result showed that BB genotype had one substitution mutation A/G in 263bp. All six breeds were at Hardy-Weinberg equilibrium（P>0.05）.Except Angus, the polymorphism information contents of other breeds were moderate polymorphisms（0.25

The inhibinβA (INHBA) gene was studied as a candidate gene for the prolificacy of Jining Grey goats. According to the sequence of bovine INHBA gene, four pairs of primers were designed to detect single nucleotide polymorphisms of exon 1 and exon 2 of INHBA gene in both high fecundity breed (Jining Grey goat) and low fecundity breeds (Inner Mongolia Cashmere goat and Angora goat) by PCR-SSCP. Only the amplified products of primer P4 displayed polymorphism. Three genotypes (AA, AB and BB) were detected in Jining Grey goats and Angora goats. Two genotypes (AB and BB) were detected in Inner Mongolia Cashmere goats. Sequencing revealed a T→C mutation at 80 bp of exon 2 of INHBA gene in genotype BB in comparison to genotype AA. This mutation resulted in an amino acid change: proline→leucine. Genotype frequency of AA, AB and BB was 0.4786, 0.4143 and 0.1071 in Jining Grey goats, respectively. In Jining Grey goats, the does with genotype AA had 1.14 (P<0.001) or 0.42 (P<0.05) kids more than those with genotype BB or AB, respectively; the does with genotype AB had 0.72(P<0.05) kids more than those with genotype BB.

The acute toxicity of heavy metal Cd to earthworm (Eisenia fetida) was studied by the method of artificial soil test. It was shown that Eisenia fetida is tolerance specie which is applicable for accumulation indication. A cDNA library of Eisenia fetida was constructed .Total RNA was extracted from Eisenia fetida after induction with Cd2+ by Trizol regent. The full-long cDNA library was constructed successfully by using the SMART technique. The results showed that the primary titer of the constructed cDNA library was 3.02×105 pfu/mL, while that of the amplified library was 8.67×109 pfu/mL .The recombination rate was about 97.3%，and the length of most cDNA inserted in the library was about 300-3000bp.The library must be helpful to make clear the components of Eisenia fetida and establish the base for research into heavy metal detoxification and immune mechanisms at molecular level.

Abstract: As a useful tools, SSR markers are widely used in research of wheat genetics and breeding. SSR protocol included the DNA extraction, PCR reaction, electrophoresis in PAGE and staining. For a more economical, simple and precise protocol of wheat SSR analysis, this paper (1) compared three methods of DNA extraction from wheat seedling, seed and embryo; (2) compared the volume of PCR mixture; (3) compared three methods of PAGE staining. A more simple SSR protocol was set up as follows: the template DNA was obtained from embryo using simplified CTAB method; 10µL PCR mixture was used instead of 20 µL; the staining protocol was only three steps by combined fixing and staining to the first step and combined developing and stopping to the second step. The simplified SSR protocol saved 50% time and 50% money than before.

The Dmrt genes, which share a highly conserved DM domain, are a new transcription factors with zinc finger domain. Although several members of Dmrt genes had been cloned from a lot of species, none was reported in shellfish. In this study, three different DM sequences（pmDmrt2，pmDmrt3 pmDmrt4） approximately 141 bp were amplified from genomic DNA of Pinctada matensii using the DM degenerate primers.The result indicate the unexpected complexity of the DM domain gene family in Pinctada matensii. The identity between pmDmrt2 and the Dmrt2 of Danio rerio, Xenopus Laevis , Gallus gallus , Mus musculu ,Homo sapiens is 95%, between pmDmrt3 and the Dmrt3 of Danio rerio, Oryzias latipes, Gallus gallus, Rattus norvegicus,Homo sapiens is 85%, and 97% between pmDmrt4 and the Dmrt4 of Oryzias latipes and Takifugurubripes. Our results further reveal the evolutionary conservation of the DM domain gene family in both invertebrates and vertebrates.

SRAP is a new molecular marker which aimed for the amplification of open reading frames(ORFs). It is simplicity, reliability, moderate throughput ratio and has not the specie-specific character. It had been widely used for gene tagging, germplasm collection,genetic diversity,map construction and comparing genome analysis. Factors influencing SRAP-PCR analysis were studied using three pear cultivars. A reliable,effective and reproductive PCR reaction system for detecting SRAP was developed. Each 20μL PCR reaction mixture consisted of template DNA 30ng, 2.0mmol/L of MgCl2, 200μmol/L of dNTPs, 30ng of primer and 1 unit of Taq polymerase.

Isoflavonoids are a subclass of plant flavonoid metabolites exclusively found in legumes, where they are important compounds mediating multiple plant-microbial interactions. The multiple health-promoting effects of isoflavonoids, especially those of typical soybean isoflavones genistein and daidzein against hormone-related cancers, osteoporosis, menopausal symptoms, and cardiovascular disease have been studied intensively. Isoflavones are secondary metabolites of plant phenylpraponoids pathway, and cloning of genes encoding IFS from several legumes has offered the possibility of genetic engineering to synthesize isoflavones in plants that normally do not yield such metabolites. However, genetic engineering studies have unveiled an integration of complex molecular regulations of isoflavones metabolic pathway.