用M13DNA探针检测小鼠DNA指纹

用ＨｉｎｆＩ酶切，随机引物法标记的Ｍ１３ｍｐ１８ＤＮＡ作探针，对无关小鼠个体及一个小鼠家系三代进行了ＤＮＡ指纹分析，无关个体平均每个个体得到１４条谱带（＞４ｋｂ）。无关个体具有相同指纹图概率为９×１０￣（－８），显示高度个体材异性，对小鼠家系内后代与亲本图谱进行了比较，表明图谱特征按孟德尔方式遗传。对家系内个体间遗传相似度Ｄ进行了计算，亲缘关系近则Ｄ值高，据此对本技术在动物个体识别，亲缘关系分析等方面的应用进行了讨论。

Morphological characteristics and DNA barcoding of Pampus echinogaster(Basilewsky, 1855)

The morphological similarities of Pampus fishes have led to considerable confusion in species-level identification,and no accurate information on neotype or DNA barcoding of Pampus echinogaster is available. Two hundred and seven specimens of P. echinogaster were collected from the coastal waters of Dandong, Dongying, Qingdao,Nantong, Zhoushan, Wenzhou, Changle, Taiwan, and Wakayama（Japan）, from June 2010 to April 2013. The diagnostic characteristics of P. echinogaster are as follows： dorsal fin VIII-XI-43–51, anal fin V-VIII-43–49, pectoral fin 22–27, caudal fin 19–22, pelvic fin absent; first gill rakers sparse, slender（pointed）, 3–4＋12–16=15–20; vertebrae39–41; transverse occipital canal on top of head moderately small, wavy ridges not reaching upper origin of pectoral fin; ventral branch of lateral line canal spare, shorter than dorsal branch of lateral line canal. By combining congener sequences of the cytochrome oxidase I（COI） gene from Gen Bank, two absolute groups were detected among all specimens, which further indicated that two valid species were present based on genetic differences in amino acid sequences and the distance between the groups. The sequences of Group 1 can be regarded as DNA barcoding of P. echinogaster. The correct morphological redescription and DNA barcoding of P.echinogaster are presented here to provide a guarantee for efficient and accurate studies, a theoretical basis for classification, and enable appropriate fishery management and conservation strategies for the genus Pampus in the future.

The Genetic Structure and Diversity of Repomucenus curvicornis Inhabiting Liaoning Coast Based on Mitochondrial COⅠ Gene and Control Region

[Object] This study was conducted to explore the genetic diversity and structure of the wild Repomucenus curvicornis inhabiting Liaoning Coast, China. [Method] The mitochondrial COⅠ gene and control region（CR） were PCR amplified from the wild R. curvicornis populations from the Liaodong Bay（n=22） and the northern Yellow Sea（n=18）, sequenced and analyzed for genetic diversity. [Result] The contents of A, T, C and G of 624 bp COⅠ gene were 24.09%, 31.04%, 25.28%, and 19.59%, and those of 460 bp CR fragment were 32.96%, 32.80%, 14.86% and 19.38%, respectively. The total number of variable sites, average number of nucleotide differences（ k）, haplotype diversity（H） and nucleotide diversity（π） based on COⅠ gene were 38, 4.67,（0.96±0.02） and（0.007 5±0.004 2）, and those based on CR fragment were 26, 3.35,（0.97 ±0.02） and（0.007 3±0.004 3）, respectively. Based on mitochondrial COⅠ gene and CR, the genetic diversity of Liaodong Bay population was lower than that of the northern Yellow Sea population. The AMOVA analysis based on CR fragments revealed almost significant genetic divergence between the Liaodong Bay and the northern Yellow Sea populations, while there was no significant genetic divergence based on COⅠ gene. The results showed that CR and COⅠ gene are effective molecular markers for detecting the genetic diversity of R. curvicornis population, while CR is more reliable than COⅠ gene in detecting the genetic structure. [Conclusion] CR is an appropriate marker for genetic analysis of marine fish population.

Immunity induced by DNA vaccine of plasmid encoding the rhoptry protein 1 gene combined with the genetic adjuvant of pcIFN-γ against Toxoplasma gondii in mice

Objective To construct the eukaryotic expression recombinant plasmid, pcIFN γ, as a genetic adjuvant and observe the immune responses elicited by pcDNA3 rhoptry protein 1 (pc ROP1) combined with pcIFN γ against Toxoplasma gondii (T gondii) infection in mice Methods A fragment of the IFN γ gene was directly inserted into the pcDNA3 plasmid and identified by two restriction endonucleases digestion pcIFN and pcROP1 DNA was injected into the left leg muscle of mice at a dosage of 100?μg, and a booster vaccination was given at the same dosage after two weeks Control groups were injected with pcDNA3 blank plasmid or normal saline At 30, 50 and 70 days after booster injection, kinetic tests were carried out: MTT assay for the proliferation response of T lymphocyte cells and the activity of NK cells, sandwich ABC ELISA for the determination of IFN γ, IL 2 and IL 10; a serum enzymetic aassay for nitric oxide (NO) in sera and ELISA for the titer of IgG antibody in sera Results The recombinant plasmid, pcIFN γ was constructed The proliferation response of spleen T lymph cells, NK cell killing activity, and serum levels of IFN γ, IL 2 and NO in mice injected with pcROP1 and pcIFN γ were higher than in those injected with pcROP1 alone There was no difference in IgG antibody levels between the two groups Conclusion The genetic adjuvant, pcIFN γ, could enhance the cellular immune response induced by DNA vaccine of pcROP1 in mice against Toxoplasma gondii infection

Crossing the DNA Rubicon

Chinese scientists make history by editing humangenes with new technologyIs it possible for humans to wall off a branch of knowledge that is deemed too dangerous,risky or formidable to pursue？That is what James Burke,the science historian,asked in his 1985 documentary series,

Inhibitory effect of retroviral vector containing anti-sense Smad_4 gene on Ito cell line, LI90

Background Transforming growth factor-β1 (TGF-β1) exerts strong fibrogenic potential in culture-activated HSCs Smad 4 is a key intracellular mediator for the transforming growth factor-β (TGF-β) superfamily of growth factors The aim of this study was to assess the effects of the antisense Smad 4 gene on Ito cell line, LI90 Methods The recombinant retroviral vector pLXSN-Smad 4 was constructed by cloning the rat antisense Smad 4 cDNA into the retroviral vector pLXSN Retroviruses with or without the antisense gene were obtained by transfecting pLXSN-Smad 4 and pLXSN vectors into PA317 cells Human hepatic stellate cells (HSCs) LI90 were infected with these retroviruses followed by selection with G418 The expression of Smad 4 was detected by Northern and Western blots Cell biological characteristics, including cell growth curve, 3H-TdR and 3H-proline uptake by HSCs and the production of extracellular matrix were assessed Results mRNA and protein expressions of Smad 4 in LI90 cells transfected with retrovirus containing the antisense Smad 4 gene were much lower than those in LI90 cells transfected with empty vector or parental LI90 cells Cells hypoexpressing the Smad 4 gene exhibited a slower rate of growth, a lower uptake of 3H-TdR and 3H-proline ( P <0 01), and smaller production of th extracellular matrix, compared with parental LI90 cells and cells transfected with empty retrovirus Conclusions The antisense Smad 4 gene can suppress the expression of the Smad 4 gene, reduce endogenous production of Smad 4 mRNA and protein, block TGF-β1 signaling pathway, inhibit activation of Ito cells, obstruct the growth of Ito cells, decrease the production of the extracellular matrix (ECM) Our results may provide a basis for the development of antifibrotic gene therapy