2003 Vol. 24, No. 3

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To investigate the effect of retinoic acid (RA),nerve growth factor (NGF),epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF) on the differentiation and apoptosis of rhesus monkey embryonic stem (rES) cell line R366.4,rES cells were cultured in monolayer state in vitro,and the experiment resulted in:1)The percentage of differentiated rES cells increased along with the elongated duration of culture.2)The rES cells differentiated into heterogeneous somatic cells,whose proportions were at random,in the absence of exogenous growth factors.3)Single growth factor could promote progress of differentiation,and increased the proportion of one or several types of differentiated cells;neuron-like cells increased in the presence of RA or NGF,while endothelium-like cells and fibroblast-like cells increased in the presence of EGF and bFGF,respectively.4)During the differentiation of rES cells the early and the late apoptosis happened,and the apoptosis could be promoted by RA and NGF.The four growth factors have the potential to stimulate differentiation of rES cells in different ways.The ES cell differentiation induced by the growth factor may be species-specific.
The pea leafminer Liriomyza huidobrensis is an invasive species,which has widely distributed in China.To specify its adaptation to cold (stress),we sampled pupae and larvae of the species from Kunming,Chengdu,Xian,Beijing and Chifeng,stepping up about every 5° from south to north.Supercooling points (SCP) of pupae were experimentally tested.The SCP increased with latitude,and the difference between populations at the two extremes was 3.7℃,and the SCP was negatively related to the yearly margin of ambient temperature for the geographical populations,but not to the monthly or yearly temperatures they experienced.The results imply that an adaptive course of lowing SCP involves in the species dispersing northwards.
A full-length cDNA of Ran gene was cloned from color crucian carp (Carassius auratus color variety) by using SMART cDNA synthesis and RACE-PCR with a pair of degenerate primers designed according to the conserved region sequence of Ran gene.The Ran cDNA was found to be 1081 bp in length with a 67 bp 5 UTR and a 366 bp 3 UTR.The coding region included 648 nucleotide acids and encoded 215 amino acids.Searching homologous genes by using this nucleotide sequence in NCBI database showed that the deduced amino acids sequence of Ran gene of color crucian carp shared high identity with Ran genes of Dano rerio (98%) and Salmo salar (97%).Moreover,the full-length sequence of its coding region was expressed in E.coli (DE[3]).To acquire multiclonal antibody,the expressed protein was purified and applied to immunize rabbits.Western blotting results indicated that the multiclonal antibody prepared by us was highly specific to recognize Ran proteins expressed in E.coli or that from eggs of color crucian carp.In addition,analysis on tissue expression specificity indicated that Ran protein was expressed in ovary,testis and kidney,whereas was not detected in heart,brain,liver,spleen and muscle.The present studies will facilitate our future studies on the physiological functions of Ran gene,the isolation and identification of its binding proteins in fish by using immunodepletion,coimmunoprecipitation,and simulative systems in vitro.
Reverse transcriptase polymerase chain reaction (RT-PCR) was employed to amplify cDNAs constructed from Trimeresurus stejnegeri venom glands poly(A)+ RNA to facilitate the cloning and sequencing of serine protease genes.The PCR products were then subcloned into pGEM-T vector and transformed into E.coli strain JM109.Five serine protease cDNAs (named TSSP-1,-2,-3,-4 and -5,respectively) were cloned and sequenced.The protease encoded by TSSP-1 is characterized by a His[41]-Arg[41] mutation in the catalytic triad.Combined the results of N-terminal sequence determinations of the purified proteins,it is concluded that TSSP-2,-3 and -4 encode stejnobin (thrombin-like enzyme),and stejnefibrase 1 and 2 (fibrinolytic enzymes),respectively.Mature proteases encoded by TSSP-1 and -2 are composed of 236 amino acid residues,and those proteases encoded by TSSP-3,-4 and -5 are all composed of 234 amino acid residues.The cloned 5 mature proteases contain 1 to 6 N-type glycosylation sites,indicating that the differences among calculated molecular weights and apparent molecular weights determined by SDS-PAGE are caused by carbohydrate content variations.Sequence identities among the cloned proteases are around 60%-90%.The differences in their structures resulted in their various substrate specificities.
For detecting the effect of population density and food availability on the growth and cannibalism,newborns of larval salamanders (Hynobius amjiensis) were basin-raised in 2×2 combinations of the ecological factors.The experiment resulted in that the somatic growth and its variation were significantly influenced by food availability,and intra-specific bite and cannibalism were determined by both the density and food availability.Of the 4 treatments,the highest growth index appeared in the combination of low-density and high-food availability,and the largest size-variation and cannibalism in that of high-density and low-food.The results were in keeping with the expect of current theories.
Using Chlorella-containing Paramecium bursaria (CCP),recovered Chlorella-containing P.bursaria (RCCP) and Chlorella-free P.bursaria (CFP) as materials,the effect of symbiotic Chlorella on the cell size,growth,resistance to oxidation,and saccharide content of the host cells were measured.The highest density of cells in the three cultures was 1500,800,500 cells/mL,respectively,corresponding to the peak period of growth on day 14,14 and 8.CCP cells averaged 22 μm longer and 16 μm wider than CFP ones,and those of RCCP cells fell between CCP and CFP.The resistance to O[2]-chemiluminescence produced by self-oxidation of pyrogallol was different:when concentrations of the cytoplasm homogenate increased from 20 to 100 μL/mL,CCP increased for 67%,RCCP for 45%,CFP for 27%,and the symbiotic Chlorella for <10%.Of glucose and maltose in cytoplasm,CCP contents were 4 and 2 times higher than those in RCCP,respectively,and CFP were only 1/4 and 1/2 of RCCP.It is very likely that paramecium-hosts benefit from the symbiotic system.
Eggs laid by migratory locusts reared under conditions of long photoperiod (LP)(L∶D=14∶10) and short photoperiod (SP) (L∶D=10∶14) were collected.Supercooling points of some eggs were measured using a thermocouple method.The remained eggs were treated with a series of low temperatures (0,-5,-10,-15 and -20 ℃) in different periods of time (6 h,1,3,5 and 10 d),and then incubated at 28 ℃.The number of surviving eggs could be judged with that of incubated nymphs,and then the survival ratio of eggs in low temperature (SL) and low-lethal temperature causing death of 50% of the eggs (LT[50]) were calculated.No difference was found in the supercooling points between LP and SP groups;SL decreased with the decline of treated temperature and the increase of treated time and SL in SP group was higher than that in LP group at -5 and -10 ℃;LT[50] increased with increase of time in low temperature,and LT[50] of SP group was lower than that of LP group.The results suggest that the cold hardiness of locust eggs could be induced by a short photoperiod experienced by parents,that is,eggs produced in autumn would better overwinter.
The partial mitochondrial COⅠ gene was sequenced from 40 individuals of four Brachionus species.The sequences were compared and their molecular phylogenetic trees were constructed by UPGMA,neighbor-joining and maximum parsimony methods using B.plicatilis as outgroup.The length of sequences was 543 bp,in which frequency of A+T was 65.3%,and a considerable variation of 157 nucleotide sites (28.9%) was detected.All of three trees suggested that B.quadridentatus is relatively closer to B.urceus than other species,and then to B.leydigi,last to B.calyciflorus.This result is consistent with that of traditional morphological taxonomy.
Tree shrew (Tupaia belangeri) is not infected by HIV-1,and its cells apparently support postentry events in HIV-1 infection.Cyclin T1 binding with HIV-1 Tat is responsible for HIV-1 transcription.To understand whether tree shrew cyclin T1 (tsCycT1) binds with HIV-1 Tat protein,the designed primers and RT-PCR were used to amplify the tsCycT1 cDNA.The clone of full-length tsCycT1 cDNA is of 2 175 bp.The homology of tsCycT1 with that of human CycT1 (hCycT1) is 92.6% at nucleotide level and 94.1% at deduced amino acid level,respectively.The first 272 amino acids have 98.8% of homology with hCycT1,and the residue at position C261 is the cysteine.These results suggest that tsCycT1 can form a high-affinity and zinc-dependent complex binding to HIV-1 Tat and TAR,which may support HIV-1 transcription.
Fourteen morphological features,and other two traits (the number of hosts,and whether or not parasitic in soft scales) were selected as characters of in-group,those of Microterys clauseni as out-group,a cladistic analysis of phylogenetic relationships among six Anicetus species parasitic on pink wax scales was made with PAUP (version 4.0b10).This analysis resulted in a tree of,that is,(A.annulatus,((A.rubensi,(A.beneficus,A.rarisetus)),(A.howardi,A.ohgushii))).It has following parameters:length=28,consistency index=0.6786,retention index=0.4375.That A.annulatus is far away from other 5 species is in keeping with the traditional observation of taxonomy.
In order to understand the molecular mechanism of the pesticide resistance of diamondback moth,Plutella xylostella,we used the SMART (switching mechanism at 5 end of RNA transcript) technology to construct full-length cDNA libraries.The PowerScript reverse transcriptase was used to synthesize and anchor the first-strand cDNA.Following reverse transcription,LD-PCR was performed using a modified oligo (dT) and an anchor primer to enrich the cDNA population for full-length sequences.The unamplified cDNA library contained 2.6×10[6] independent clones,in which the recombinant clones with an average insert size of 1.6 kb was > 96%.The titer of the amplified library was 2.4×10[10] pfu/mL.PCR analysis of 60 randomly selected clones showed that 65% of cDNA clones contained inserts of above 1.0 kb.From the cDNA library,a full-length ribosomal protein L27A cDNA with 5 and 3 untranslated regions was isolated.
The morphology and fine structure of the compound eye in Scylla serrata was observed with electron microscopes (EM).Under the scanning EM,there was a thumb-shaped area with no ommatidia on the back of the compound eye.Under the transmission EM,ommatidia were hexagon,which are specific to Decapoda Brachyura;the sensitization system of an ommatidium included 11 retinular cells (RCs),of which four located at the distal part of the ommatidium;seven formed the proximal main part of the retina.Thus a "4+7"structure could be seen at the conjunction area,partly in overlap.These observations are in keeping with those reported for Portunus tritubereulatus and the larval of Macrobrachium rosenbergii,but different from those with the structure of "1+7" in Libinia and Eriocher sinensis.The difference may be related to the dependence of the species on the visual sense in their natural habitats.
Four new species of Lordiphosa denticeps species-group are reported from Yunnan,China,namely Lordiphosa gruicollara sp.nov.,Lordiphosa eminens sp.nov.,Lordiphosa incidens sp.nov and Lordiphosa piliferous sp.nov.The distribution of the species of denticeps species-group shows distinct geographical replacement.
The success of different stages of reproduction in cormorants (Phalacrocorax carbo) was followed at Luci Island in Qinghai Lake in the breeding season from March to August 2000.The clutch size was 3.5;hatch rate,survival rate of nestlings,and total rate of reproductive success was 37.9%,61.1%,and 23.2%,respectively.The loss rate was 36.1% for nests in first 3 days of incubation,and 25% for nestlings in the first 10 days.These data may be viewed as part of the baseline for long-term monitoring changes in the bird population,which is an important element of the wetland ecosystem.
Experimental mouse were treated with three living densities,2,6,and 10 individuals per cage (26 cm×15 cm×15 cm).After a week of the treatment,livers of the mouse were collected for determining the content of lipid peroxides,and the activities of the catalase and the superoxide dismutases with thiobarbituric acid reaction,colorimetry and nitrite-form method,respectively.The content of lipid peroxides in 6 ind.-group and 10 ind.-group was significantly higher than that in 2 ind.-group.The activity of the catalase in 10 ind.-group was lower than that in 2 ind.-group,suggesting that crowding results in the decline of the catalase activity,which induces the increase of lipid peroxides,bringing peroxidatic damnification to the body.