Biomedical subjects

R Jiao

Publications and source records attributed to R Jiao.

13 recordsLinked to original sources

PubMed2001

Headless flies generated by developmental pathway interference.

Ectopic expression of transcription factors in eye-antennal discs of Drosophila strongly interferes with their developmental program. Early ectopic expression in embryonic discs interferes with the developmental pathway primed by Eyeless and generates headless flies, which suggests that Eyeless is necessary for initiating cell proliferation and development of both the eye and antennal disc. Interference occurs through a block in the cell cycle that for some ectopic transcription factors is overcome by D-CycE or D-Myc. Late ectopic expression in cone cell precursors interferes with their differentiation. We propose that this developmental pathway interference is a general surveillance mechanism that eliminates most aberrations in the genetic program during development and evolution, and thus seriously restricts the pathways that evolution may take.

Animals↗

PubMed2001

[Studies on regulation of glutamine synthetase activity from Streptomyces lincolnensis].

Glutamine synthetase in crude extracts from Streptomyces lincolnensis growing under different nitrogen sources were studied. The results showed that NH4+ in high concentration repressed the biosynthesis of the enzyme. To determine whether Streptomyces lincolnensis has undergone covalent modification, a comparison of the glutamine synthetase isolated from cells grown on different nitrogen sources was made. No significant difference was observed in specific activity, pH optima, divalent cation response, and ultraviolet absorption spectra. Glutamine synthetase activity was not influenced by ammonia shock or snake venom phosphodiesterase treatment. Under these conditions, the activity of glutamine synthetase from K. aerogenes was markedly changed. There was therefore no evidence for enzymatic adenylylation of glutamine synthetase from Streptomyces lincolnensis. Glutamine synthetase was subject to feedback inhibition by end products of glutamine metabolism. Cumulative feedback inhibition of the Mn(2+)-dependent glutamine synthetase activity was demonstrated. These results suggest that glutamine synthetase from Streptomyces lincolnensis is an allosteric enzyme.

Ammonia↗

PubMed2000

[The selection of fluoroacetate-resistant mutant from E. coli MMR204 and its influence on the expression of heterologous GL-7ACA acylase].

In the cultivation of gene engineered strain of Escherichia coli on glucose medium, excretion and accumulation of acetic acid inhibit not only cell growth but also the the expression of heterologous protein. It is obvious that the desirable host strain maintaining acetate at a low level is one of the approaches to increase the production of recombinant protein. The present article deals with the selection of mutants of E. coli DP19, DP8, which grow on the medium containing pyruvate as the sole carbon source in the presence of 50 mmol/L fluoroacetic acid. It is shown that mutant DP19 is defective in its phosphotransacetylase(PTA) activity and accumulates less acetate in the medium, while DP8 is defective in acetate kinase (ACK) and accumulates similar level of acetate comparing with its parent. Using pta- mutant E. coli DP19 as host, the expression of GL-7ACA acylase gene on the recombinant plasmid pMR24 is improved, and the yield of enzyme activity in flask fermentation is about twice as much as its parent.

Acetate Kinase↗

PubMed1999

[Cloning of mitomycin C resistance gene from Streptoverticillium caespitosum ATCC27422 and studying its function].

Streptoverticillium caespitosum is the major producer of anti-cancer drug mitomycin. It demonstrated that there exists a strong restriction system for the transformation of foreign DNA in this strain. Using the blocked mutant S-6 which is sensitive to mitomycin C (MC) and does not produce mitomycin as a recepient and plasmid pIJ699 as vector, the gene library of the strain ATCC27422 was constructed. A clone of mitomycin C resistance gene(mcr) was obtained by screening from this library. The mitomycin C resistance gene cloned on the 6.2 kb fragment of plasmid pLX5 could be expressed in the Strptonmyces lividans TK54. It was the first time to introduce successfully the plasmid pLX5 into the wild type strain ATCC27422 by electroporation technique. And the transformant's resistance to mitomycin C has risen greatly: the MIC has gone up from 200 micrograms/ml to more than 1000 micrograms/ml. The result of fermentation showed that the yield of mitomycin of ATCC27422 harboring plasmid pLX5 was more than the wild strain ATCC27422's. It suggested that there is some relationship between the level of the resistance to mitomycin C and its biosynthesis.

Antibiotics, Antineoplastic↗

PubMed1999

[Conjugational actinomycetes of plasmid RSF101 from Escherichia coli to the rare actinomyceces of Nocardia asteriodes and Streptoverticillum caespitosus].

RSF1010 is a naturally occurring Escherichia coli broad host-range plasmid about 8.7 kb in size. It can be mobilized at high frequency between different gram-negative bacterial species when transfer functions are available in trans. Following the pioneering work of conjugational transfer of RSF1010 from E. coli to Streptomyces lividans and Mycobacterium smegmatis, the transfer of this plasmid by conjugation from E. coli S17.1 tp two gram-positive rare actinomycetes, Nocardia asteroides 3927 and Streptoverticillum caespitosus ATCC27422 was first time reported in this study. Southern blot analysis of the total DNA extracted from the actinomycetes' exconjugants proved that RSF1010 had been transferred from E. coli into the two new hosts and maintained staby in the exconjugants. Meanwhile, partial deletions of RSF1010 replicon loosing its antibiotics resistance makers were readily detected in E. coli. The implenmentation of this observation was discussed.

Actinomyces↗

PubMed1998

[Studies on characterization of glutamine synthetase from Streptomyces lincolnensis].

Glutamine synthetase from Streptomyces lincolnensis was purified to electrophoretic homogeneity. The effects of pH, temperature and metal ions on the purified enzyme were studied, and the substrate specificity of glutamine synthetase were reported. Under the standard biosynthetic assay system, the substrate saturation curves of glutamine synthetase from Streptomyces lincolnensis for ammonia did not agree with the Michaelis-Menten relationship, with either Mn2+ or Mg2+ as divalent cation, and Hill constants not equal to 1. The K(m) values of GS for Glu and ATP were different with different divalent cation.

Glutamate-Ammonia Ligase↗

PubMed1998

[Purification and properties of alanine dehydrogenase from Streptomyces lincolnensis].

Alanine Dehydrogenase (L-Alanine: NAD+ oxidoreductase, deaminating, EC 1.4.1.1) was purified from Streptomyces lincolnensis through four steps: (NH4)2SO4 precipitation, DEAE-cellulose 52, Affi-Gel Blue and Sepharose 6B. Molecular weight of the enzyme was determined as 170,000 by gel filtration and concentration gradient PAGE. SDS-PAGE showed only one band of 42,500, demonstrating that ADH from Streptomyces lincolnensis was consisted of four identical subunits. The optimal pH for amination was 9.0, for deamination 9.5. The optimal temperature for both amination and deamination was 50 degrees C. The Km valuse for pyruvate, NH4+, NADH, L-Ala and NAD+ were 2.08 x 10(-4) mol/L, 2.00 x 10(-2) mol/L, 2.38 x 10(-5) mol/L, 1.43 x 10(-2) mol/L and 6.67 x 10(-5) mol/L, respectively.

Alanine Dehydrogenase↗

PubMed1996

Signal transduction pathways in guinea pig sperm.

Trifluoperazine (TFP), the antagonist of calmodulin (CaM), significantly stimulated the capacitation and acrosome reaction of guinea pig spermatozoa at the concentration of 10-100 mumol/L, independent of the external Ca2+. Forskolin, dbcAMP and caffeine evidently promoted the occurrence of acrosome reaction of spermatozoa at early capacitation stage (5 h) in nonsynchronous system but not in synchronous system. If the spermatozoa were capacitated for 15 h in synchronous system, the above three drugs significantly stimulated acrosome reaction in a Ca(2+)-independent manner. Protein kinase C activators, i.e. phorbol 12-myristate 13-acetate (PMA) and phorbol 12,13-dibutyrate (PDB) did not influence the occurrence of acrosome reaction of spermatozoa at early capacitation stage, but significantly increased the acrosome reaction rate in capacitated spermatozoa in a Ca(2+)-independent manner. In contrast, PKC inhibitor staurosporine significantly inhibited the occurrence of acrosome reaction.

Acrosome↗

PubMed1994

Nitrite reductase gene cloning of Amycolatopsis mediterranei U-32.

Southern blot analysis showed great homology existed between niaD (NR gene) of Aspergillus nidulans and A. mediterranei U-32 chromosome DNA. A 5.0kb PstI fragment from A. mediterranei U-32 complementary to A. nidulans niaD gene was cloned in E. coli NM522 using niaD as a probe. An identical DNA band was observed through back-hybridization of the cloned DNA fragment to PstI digest of A. mediterranei U-32 chromosome DNA. Its 2.1 kb SmaI-EcoRV fragment can only hybridize with total RNA from nitrate-cultured mycelium but not with that from ammonia-cultured mycelium. These data suggested that the cloned DNA fragment contains NR gene of A. mediterranei U-32. This is the first report on NR gene cloning from a aerobic bacterium. It was deduced from the molecular weight of the nitrate reductase that the coding sequence of NR gene is almost 1.5 kb in size. Further hybridization analysis indicated that the cloned DNA fragment covers the full-length NR gene of A. mediterranei U-32. We also constructed the physical map of the recombinant plasmid pJL1 with various restriction endonucleases, among them ten with no restriction site, six with unique site and two with double sites on the insert.

Actinobacteria↗

PubMed1992

Localization of adenovirus DNA by in situ hybridization electron microscopy.

Biotinylated deoxyadenosine triphosphate (dATP) (Bio-7-dATP) and 3H deoxythymidine triphosphate (dTTP) labeled adenovirus DNA were hybridized in situ to thin sections of Lowicryl K4M-embedded and whole-mount extracted HeLa cells infected with adenovirus. The biotinylated probe was detected by exposing the extracted cells or sections to antibodies against biotin followed by colloidal gold-conjugated secondary antibodies and then critical-point dried while 3H-dTTP labeled probe by electron microscopic autoradiography. On Lowicryl K4M sections, gold particles and silver grains were mainly restricted in the nucleus. Furthermore, whole-mount results suggested that replicating adenovirus DNA is localized on the nuclear matrix of its host cell. In this paper, the described non-radioactive procedures for hybrid detection offered several advantages: a) rapid signal detection; b) superior morphological preservation and spatial resolution; c) precise localization; and d) on Lowicryl K4M sections, signal to noise equivalent to radiolabeling.

Acrylic Resins↗

PubMed1992

Restriction mapping and localization of GL-7-ACA acylase gene.

This paper presents the results about the restriction mapping of recombinant plasmids pMR5 and pMR6 containing GL-7-ACA acylase gene from Pseudomonas sp. 130, gene localization and its expression under the control of different promoters, tet, tac or lac/tac, in Escherichia coli. The analysis of gel electrophoresis of pMR5 cleaved with several kinds of restriction enzymes indicated that there is no sites of EcoRI, HindIII and ClaI but the presence of following sites: one HpaI, two XhoI, three EamHI and four PstI on the cloned gene fragment. The restriction maps of pMR5 and pMR6 were determined by comparative digestion of various endonucleases. The gene of GL-7-ACA acylase was localized on a 3.0kb fragment of B2-B3-HpaI from the studies on a serial subcloning. Expression of subclones pMR9, pMR10 and pMR11 in E. coli was compared. Higher yield of acylase was obtained when the gene fragment was placed downstream of the tac promoter. The expression of Pseudomonas gene in E. coli was also discussed.

Amidohydrolases↗

PubMed1991

[The study of Bacillus sphaericus Ts-1 protoplast-plasmid transformation of electroporation].

This report gave the best conditions of Bacillus sphaericus Ts-1 protoplast-plasmid pHV33 electroporation. The highest transformation frequency and transformation efficiency induced by three pulse of 21 KV/cm and 10 microseconds duration applied at an interval of one sec., was 2.44 x 10(2) transformants/micrograms DNA and 3.16 x 10(-6) respectively. The saturated concentration of DNA absorbed by the protoplast was 5 micrograms DNA/10(9) cells/ml. By means of this method, pJB417, a recombinant mosquito larvicide clone, was introduced into B. subtilis 168M and B. sphaericus Ts-1. The transformants of B. subtilis 168M with biocide activity were obtained, but the toxicity of B. sphaericus Ts-1 was not increased.

Animals↗

PubMed1990

[The relationship of Sindbis virus assembly and the viral protein 6K with intermediate filaments].

The relationship of Sindbis virus (Sbv) assembly with intermediate filaments was studied by means of whole-mount and DGD embedment-free technique of EM together with the procedure of gentle extraction. In the early stage of Sbv infection, the "virus assembly center" was suspended in the intermediate filament network. In the late stage, the assembling and assembled virus nucleocapsids were associated with intermediate filaments. It is presumable that the virus nucleocapsids may move from the "virus assembly center" to cytoplasmic membrane along the intermediate filaments. Further study using immunolabelling technique indicated that the nonstructural protein 6K of Sbv was probably associated with intermediate filaments as well.

Cell Line↗