ALTERATION IN CHROMATIN CONFORMATION OF PROLIFERATING AND DIFFERENTIATING ENZYME GENES IN NORMAL MOUSE LIVER AND ASCITES HEPATOMA CELLS AND ITS RELATION TO THE GENE EXPRESSION OF ENZYMES

Nuclei from the normal mouse liver were partially digested with micrococcal nuclease, followed by DNA extraction, agarose gel clectrophoresis and dot blot hybridization with ~(32)P-labeled cDNA probes of CPS_1 and ACT complex, It was clearly shown that the CPS_1 genes were distributed on the monomer, dimer. and trimer of nucleosomes, while the genes coding for ACT complex were distributed on the condensed oligonucleosomes. An opposite manner of distribution of CPS_1 and ACT complex genes was, however, noted in the case of ascites hepatoma cells, in which the specific activity of ACT was 13 times higher than that in the normal liver, while that of CPS_1 was remarkably reduced. Similar patterns of change in mRNA level of CPS_1 and ACT complex were observed in the normal mouse liver and ascites hepatoma cells, indicating a close relationship between chromatin structure and gene expression of these enzymes.     

MOLECULAR CLONING AND EXPRESSION OF Bacillus thuringiensis subsp. galleriae INSECTICIDAL CRYSTAL PROTEIN GENES IN Escherichia coli

The location of the toxin gene of B. thuringiensis subsp. galleriae (H5ab) on the Mr-130Mdplasmid is determined by molecular cloning. Double digestion fragments (BamHⅠ and SalⅠ)and PstⅠ restriction fragments as well, from the 130 Md plasmid of B. thuringiensis subsp.galleriae, are ligated with the cloning vector pAT 153 respectively and transformed into E.coli strain HB 101. Out of 208 transformants, three colonies (FG2, FG9, FG19) give posi-tive hybridization reaction using the HD-1 delta-endotoxin gene as a probe. They are presum-ed to contain the delta-endotoxin gene of B. thuringiensis subsp. galleriae. Western bolt assaysindicate that Mr-130 kDal and 68 kDal, crystal proteins produced by clone FG 2 react withanticrystal protein antibody. The protein extracts of clone FG2 are lethal to Ostrinia furna-calis (Guenee). This is the first report with regard to the cloning and expression of the B. thuringiensissubsp. galleriae (H5ab) delta-endotoxin gene.     

INTRODUCTION, EXPRESSION AND REGENE-RATION OF THE TiT-DNA GENES IN CULTURED CELLS FROM Hyoscyamus muticus +Nicotiana tabacum SOMATIC HYBRIDS

After the cultured cells from Hyoscyamus muticus + Nicotiana tabacum somatic hybridswere cocultivated with different virulent strains of Agrobacterium tumefaciens harboringoctopine- type Ti plasmid or nopaline- type Ti plasmid using the transformation procedurein vitro developed in the present investigation, the TiT- DNA genes were introducedinto the host cells. The onc genes and ocs or nos genes located on TiT- DNA were expres-sed in transformed colonies derived from the cocultivated cells. Although the platingefficiencies of recipient cells were reduced by the agrobacterial treatment, the frequenciesof phytohormone autotrophy ranged from 33.9 to 76 .8% in the cells infected with viru-lent strains in hormone- free conditions, and the frequencies of opine synthase activityamounted to 9.7- 47 .5%. Teratomatous shoots were regenerated from the transformed col-onies. During the course of culture the shoots were no longer to lengthen when theygrew up to 1 -3 cm in length, and they could not be rooted. Follo     

INTRODUCTION OF FOREIGN GENES INTO THE SEED EMBRYO CELLS OF RICE BY ELECTRO-INJECTION AND THE REGENERATION OF TRANSGENIC RICE PLANTS

The NPTII gene has been successfully transferred to the seed embryo cells of two rice varieties (Oryza sativa L. subsp. indica cv. Sanerai and Qryza sativa L. subsp. japonica cv. Nonghu No. 6) by means of electroinjection. Resistant calli were screened out on MS medium with 100 μg/ml Km. Transgenic rice plants were regenerated via somatic embryogenesis. Both NPTII detection and Southern blot hybridization demonstrate that the foreign gene has integrated and expressed stably in the transformants.     

TRANSFER OF THE ATRAZINE-RESISTANT GENE OF BLACK NIGHTSHADE TO SOYBEAN CHLOROPLAST GENOME AND ITS EXPRESSION IN TRANSGENIC PLANTS

The atrazine-resistant psbA gene of black nightshade was transferred to the chloroplast genome of atrazine-susceptible soybean by means of ovary microinjection during the stage of zygote. The identification was carried out by using the methods of spraying the leaves directly with atrazine solution, examining the change of leaf fluorescence kinetics under a brighter light induction, molecular hybridization, etc. The experimental results show that the transgenic soybean plants do have been obtained for the first time.     

EXPRESSION OF SYNTHESIZED HIRUDIN GENE IN YEAST

Hirudin is a sort of polypeptides secreted from the salivary gland of medicinal leech. It is of potential importance in medicine. We designed and synthesized the hirudin gene based on the amino acid sequence of hirudin HV2, and expressed it using the yeast alpha factor expression system. The yeast strain stably carrying the hirudin expression plasmid was deduced by mutagenesis. After its cultivation in rich nutritious medium for 36—48 h, the hirudin expression product secreted into the culture fluid was 10—20 ATU/ml. The HPLCpure hirudin product could be obtained through a simpler purification procedure, its N-terminal amino acid sequence was identical with the natural product, and it showed potent anticoagulant and antithrombin activity. About 3000 ATU of pure hirudin witha specific activity of 6600 ATU/mg could be obtained from 500 ml of culture fluid.     

CLONING, EXPRESSION AND CHARACTERIZATION OF Pro3 GENE OF YEAST Saccharomyces cerevisiae

The proline biosynthetic pathway and Pro genes in Saccharomyces cerevisiae have just begun to be studied recently. In our laboratory, Pro2 gene of S. cerevisiae had been cloned in yeast. As described in this paper, yeast Pro3 gene was also cloned, which can complement yeast Pro3 mutants, and be expressed efficiently in E. coli. The high activities of this gene product, L-pyrroline-5-carboxylate (P5C) reductase, can be detected in both organisms. The activity of the Pro3 gene product in multiple copy plasmids is not higher than that of single copy genes in chromosomes in both yeast and E. coll. The preliminary characterization of the gene is also reported.     

RETROVIRUS MEDIATED TRANSFER OF ANTISENSE HUMAN c-myc GENE INTO HUMAN ESOPHAGEAL CANCER CELLS SUPPRESSED CELL PROLIFERATION AND MALIGNANCY

A retroviral vector, called pDAM3, containing the neomycin resistant gene and the antisense human c-myc gene fragment (the third exon and 3'flanking sequence) was constructed. pDAM3 was introduced into amphotropic packaging cells PA317 by the calcium phosphate precipitation method. Several G418-resistant PA317 clones were isolated. The virus titer of these cell lines was determined by infectivity of their cuhure fluid to NIH/3T3 cells. The highest titer obtained was 8×10~5 G418-resistant colony forming units/ml. Clonal and pooled G418-resistant PA317 colonies with high titers were expanded and analyzed by Southern blot for the presence of intact viral sequences. All cell lines were found to harbor the internal sequences of the pDAM3 vector without any rearrangement. Recombinant virus DAM3 infected human esophageal cancer cell line EC8712 efficiently. The DAM3-infected EC8712 (called EC-DAM3) was found to contain the full DAM3 sequence (4.8kb) by Southern blot analysis. Antisense myc RNA expressed in th     

STUDIES ON INTRODUCTION OF FOREIGN GENES INTO CULTURED CELLS OF Oryza sativa INDICA USINC Agrobacterium Ti PLASMID SYSTEM

Either bacterial attachment or cellulose fibrillar elaboration was hardly observedduring the cocultivation of the cultured suspension cells of Oryza sativa Indica with thestrain C58C1 Rif~r of Agrobacterium tumefaciens that was not specially pretreated. Onthe other hand, quite a lot of Agrobacterium cells were found to adhere to the surface ofcultured rice cells and a number of cellulose fibrils were produced around the specifiedbacteria when phenolics-pretreated bacteria were cocultivated with rice suspension cellsin common culture media, especially in complex culture solutions. The complex culturesolution was the bacterium-free filtrate of hormone-containing MS medium which hadbeen utilized to incubate carrot cells and the newly wounded hypocotyl segments fromtomato and Agrobacterium cells. Detecting experiments demonstrated that both NPT Ⅱ andNOS genes, located on the T-DNA segment of chimaeric plasmid pGV3850 :: 1103neo, weretransferred and expressed in the cultured cells of O. sativa Indica in the     

DEVELOPMENT AND IDENTIFICATION OF CHINESE SPRING Ta1 kr ph1b PLANTS AND THEIR USE ON GENETIC TRANSFER OF DESIRABLE GENES FROM Aegilops

Chinese Spring Tal kr phlb plants were developed by hybridizing Tal common wheat and Chinese Spring phlb mutant and backcrosses. When Chinese Spring Tal kr phlb plants were crossed with some wild species of Triticeae, much more seeds of wide hybridization can be obtained without artificial emasculation and pollination because of the existence of Tat and kr genes. The homoeologous chromosomes between common wheat and the wild species of Triticeae can also pair and exhibit cross-over in the hybrid F1 plants because of the existence of the phlb gene. So that more alien desirable genes can be transferred to common wheat rapidly. Therefore, Chinese Spring Tal kr phlb plants become a powerful tool for wide hybridization and transferring alien desirable genes to common wheat.     

THE ANTISENSE C-HA-RAS OLIGODEOXY-NUCLEOTIDES INHIBIT THE PROLIFERATION OF THE GASTROCARCINOMA-DNA-TRANSFORMED RAT 3-3 CELLS AND REDUCE THE P21 EXPRESSION OF THE CELLS

The Rat 3-3 is a secondary transformant of the rat fibroblast cell line (Rat-1) transfectedwith total DNA of a gastrocarcinoma cell line BGC-823. The cells over-express the c-Ha-rasoncogene which contains point mutation at the 12th codon. In order to determine how theactivated c-Ha-ras oncogene expression governs the cell's transformation, two pendadecadeoxy-nucleotides AS-1 and AS-2 were synthesized. AS-1 was complementary to the single strandof the first three codons and the upstream sequence close to the ribosome binding site of c-Ha-ras mRNA. AS-2 was complementary to the 3' end of the first intron and the 5' end ofthe second exon of c-Ha-ras unripe RNA enclosed in the nucleus. The oligonucleotides couldblock either the translation of c-Ha-ras mRNA or the splicing of c-Ha-ras unripe RNA, thusinhibiting the expression of the activated c-Ha-ras oncogene and the proliferation of thetransformed cells Rat 3-3. The inhibitory effect increased with a growing concentration ofthe antisense oligodeoxynucleoti     

INTERACTION OF 8-METHOXYPSORALEN AND NEAR-UV LIGHT CAUSES MUTATION AND CYTOTOXICITY IN MAMMALIAN CELLS

Abstract The interaction of near-UV light and a photosensitizer, 8-methoxypsoralen (8-MOP), was studied in the Chinese hamster ovary cell/hypoxanthine-guanine phosphoribosyl transferase system; cell survival (cloning efficiency) and mutation induction (resistance to 6-thioguanine) were quantified. Exposure of cells to either 8-MOP up to 20 μg/m l (93 μ M ) or near–UV light up to 40000 J/m² had no effect on either survival or mutation frequency. Preincubation of cells with 8–MOP from 5 to 120 min prior to irradiation with various fluences did not affect cell survival or mutation frequency. Survival decreased and mutation frequency increased linearly when either the 8-MOP concentration or fluence was increased while the other factor was held as a constant. Mutation frequency appears to show reciprocity relative to the product of 8-MOP concentration times fluence of near–UV light [(μg/m l )·(J/m²)] throughout a range apparently limited by high cell lethality. The observed pooled data on mutation, f (x), as a function of (μg/m l )·(J/m²), x , fit a linear dose–response line, f (x) = (34.2 + 0.05 x ) × 10^-6. Cell survival, however, does not appear to exhibit such reciprocity.     

时间域曲线拟合求取糠醛加氢反应的传递,吸附与动力学参数

利用脉冲-应答技术,测定了糠醛在BHK-1催化剂上的传递与吸附系数和加氢反应速率常数。依据修正过的Kubin-Kucera气固相传质一维扩散模型,用单纯形法进行多维搜索的最优化方法来完成富里埃分析和时间域曲线拟合的计算。求得了六个参数:E、D_e、k_f、k_a、K_a、k_r,并对富里埃分析法和时间域曲线拟合法作了比较。     

基于催化链转移聚合法MMA/MPS共聚物的制备及其表观催化链转移常数测定的研究

以钴Ⅱ肟氟化硼络合物(CoBF)为催化剂,2,2′-偶氮二异丁腈(AIBN)为引发剂,实现了甲基丙烯酸甲酯(MMA)与γ-甲基丙烯酰氧基丙基三甲氧基硅烷(MPS)在60℃甲苯体系中的催化链转移聚合(catalytic chaintransfer polymerization,CCTP),制备出末端含有双键的共聚物.利用核磁共振证明了其末端双键的存在,并通过热重分析证明CCTP产物与自由基聚合产物的结构区别.用凝胶渗透色谱(GPC)对7种单体组成下不同催化剂CoBF用量的聚合产物进行分子量表征,结果表明以催化链转移聚合合成的共聚物具有分子量低及分子量分布较窄,且聚合物的分子量随着催化剂CoBF的增加呈明显下降趋势.又分别采用了基于DPn(数均聚合度)、DPw(重均聚合度)的Mayo方程和基于ΛP、ΛH的链长分布方程计算出催化剂的表观链转移常数,发现基于DPw的Mayo方程和基于ΛP的链长分布方程的计算结果最为接近.并通过对共聚体系中不同单体组成的研究发现,催化剂表观链转移常数随着单体组成中MPS的增加而增加.     

共轭聚合物/C60复合体系及其在光伏打电池中的应用

共轭聚合物/C₆₀复合体系在有机太阳能电池中的应用引起了化学工作者的广泛兴趣.本文介绍了共轭聚合物/C₆₀复合体系的光诱导电子转移,以及近年来该体系在光伏打电池中的研究进展.     

取代酚及其酚钾与四氰基乙烯的电荷转移络合作用

本文以八种取代酚及相应的取代酚钾作为电子给体,以四氰基乙烯(TCNE)作为电子受体,在乙腈溶液中用紫外可见分光光度法测定了取代酚及相应的取代酚钾与TCNE作用的电荷转移络合物的光谱,以其CT光谱用比较法计算了取代酚及相应的取代酚钾的电离势。结果表明,除4-氰基苯酚外,其余取代酚及取代酚钾均可与TCNE形成电荷转移络合物,取代酚钾比相应的取代酚有更好的给电子能力,取代酚的电子转移能与Hammet取代基常数有较好的线性相关关系。     

DNA STRAND BREAKS IN MAMMALIAN CELLS EXPOSED TO LIGHT IN THE PRESENCE OF RIBOFLAVIN AND TRYPTOPHAN

Abstract— When mammalian cells were exposed to visible-fluorescent light or near-UV light in the medium containing riboflavin and L-tryptophan, single-strand breaks appeared in their DNA. This did not occur if either riboflavin or tryptophan was omitted from the medium. The same effect was observed when cells were added to the pre-irradiated medium, indicating that a stable photoproduct was responsible. The induced DNA lesions were shown to be equally repairable in both excision proficient and defective (xeroderma pigmentosum) human cell lines. The active photoproduct formed was shown to be hydrogen peroxide. The possible relationship between these results and the near-UV induced killing of mammalian cells is discussed.     

TERMINATION AND TRANSFER OF THE CHAIN RADICALS IN THE POLYMERIZATION OF ACRYLONITRILE INITIATED BY VANADIUM(V)-THIOUREA REDOX SYSTEM

The dependence of the molecular weights on the concentration of reactants in the polymerization of acrylonitrile initiated by vanadium (V)-thiourea redox system has been investigated. It was found that the molecular weights of the polymer change nonlinearly with increasing concentrations of nitric acid and thiourea. Probably, the composition of the complexes exert a great influence on the chain initiation and termination. The reaction of "complextermination" gives rise to the decrease of the molecular weights markedly while the concentrations of thiourea and vanadium (V)in the range from one to three molar ratios.     

ELECTROFUSION OF IBRS2 CELLS AND THE STUDY OF THEIR FUSION PROCESS

IBRS2 epithelial cells in monolayer culture fused at a very high frequency when exposed to high-voltage electric pulsing fields. Exposure to four repetitive electric pulses of about 1.7 kilovolts per centimeter with a duration of 100 microseconds caused more than 90 percent of the cells to become fused (multinucleate) when 1 millimolar magnesium was present in the pulsing medium. Magnesium and calcium ions in the pulsing medium had a very strong effect on the electrofusion of IBRS2 cells. Magnesium could increase not only the electrofusion yield but also the stability of the cells under the conditions of electrofusion. In contrast, calcium inhibited electrofusion and decreased the stability of the cells. Careful microscopic observation revealed the electrofusion of IBRS2 cells to be very complex, dynamic process undergoing many interesting changes. A possible explanation for the process and mechanism of electrofusion of IBRS2 cells was proposed in agreement with the experimental observation.