运用差异展示分离特异性表达的基因

在高等生物中含有约100000个不同的基因,其中仅有15％的基因在任何个体细胞中均表达．因此分离特异的目的基因便显得十分重要.差异展示是通过部分扩增mRNA的逆转录产物、经测序胶电泳,分离到差异性表达的基因.它与消减杂交相比是分离特异表达基因的更有效的手段．虽然这种方法在实际运用中存在着这样或那样的困难,但随着对这种技术的不断改进,它将会有越来越广泛的用途．     

Luteal function in the hysterectomized bitch following treatment with prostaglandin F2α (PGF2α)

Estrus and ovulation were induced in ten mature, mixed-breed, anestrous bitches (10 to 20 kg) using exogenous gonadotropins. Bitches were bred once, on the second day of estrus. Between 11 and 13 days following estrus, bitches were bilaterally hysterectomized and randomly divided into two treatment groups of five bitches each. Four days following surgery, Group A (treated) was given a single subcutaneous injection of PGF₂α (Prostin F₂ alpha^®) at a dose of 1 mg/kg body weight and Group B (controls) similarly given an equal volume of .9% saline. Blood samples were collected daily by cephalic venipuncture prior to surgery and for 75 days thereafter. Plasma progesterone was monitored by a radioimmunoassay method. Although bitches were teased daily following PGF₂α or saline treatments, estrual behavior was not exhibited. In both the PGF₂α and saline treatment groups, plasma progesterone levels showed a transient decline by 12 hours following injection, although a more dramatic decrease was observed at this time in the prostaglandin-treated bitches. Subsequently, progesterone concentrations tended to increase in both groups at 6 days following treatment, however, not to pre-treatment levels. Within 20 to 32 days following treatment in both groups, plasma progesterone levels declined to <1 ng/ml and remained depressed at least 60 days post-injection. In this study, complete luteal regression was not induced following PGF₂α treatment. Luteal function in both groups, as indicated by plasma progesterone concentrations, was shortened in the absence of the uterus.     

Effects of taurine and γ-aminobutyric acid on akinesia and analgesia induced by D-Ala2-Met-enkephalinamide in rats

Effects of taurine or γ-aminobutyric acid (GABA) on akinesia and analgesia induced by D-Ala²-Met-enkephalinamide were investigated in rats. Administration of taurine (dose range: 2.375×10^?2 M–9.5×10^?2 M/10 μl) into the left lateral ventricle 10 min prior to the injection of D-Ala²-Met-enkephalinamide (50 μg/10 μl) produced a dose-dependent reduction in the duration of akinesia and to some extent of analgesia, as estimated at 30 min and 60 min following the enkephalinamide injection; at the first estimation-time (10 min), taurine did not alter the duration of akinesia or that of analgesia. The median effective dose (ED₅₀) for akinesia determined at 60 min after D-Ala²-Met-enkephalinamide was 5 times greater and that for analgesia assessed at the same time was 1.7 times greater in taurine-treated rats than the respective doses in control animals. Administration of GABA under similar experimental conditions produced a dose-dependent reduction in the duration of analgesia from the initial estimation time (10 min) following the injection of D-Ala²-Met-enkephalinamide. The ED₅₀ for analgesia determined at 30 min after D-Ala²-Met-enkephalinamide was 3 times greater in GABA-treated rats than in control animals. Unlike the effects of taurine, GABA did not alter the duration of akinesia. Neither the duration of akinesia nor that of analgesia was modified by taurine or GABA alone in rats tested 9 min after the injection of each amino acid. These findings suggest that taurine may promote a recovery from both akinesia and analgesia, while GABA decreases only the analgesia induced by D-Ala²-Met-enkephalinamide.     

Studies on non-H-2-linked lymphocyte-activating determinants. IV. Ontogeny of the Mls product on murine B cells.

The minor lymphocyte stimulating (Mls) locus codes for lymphocyte activating determinants (LADs) on murine B lymphocytes, but not T lymphocytes. This observation was strengthened by a series of techniques which allow deletion and addition of T and B cells. These included the use of cytotoxic antisera such as anti-Thy 1.2, anti-MTLA, anti-MBLA, and complement, and the use of a goat anti-μ antisera, and finally the use of a fluorescence activated cell sorter (FACS).The studies in this report document the organ distribution and the ontogenetic appearance of the surface LADs on the surface of B lymphocytes from DBA/2N (H-2^d, Mls^a) and CBA/J (H-2^k, Mls^d) mice. Adult-like ability to stimulate H-2 identical BALB/c (H-2^d, Mls^b) and C3H/He (H-2^k, Mls^c) responder cells appeared at about 4–5 weeks of age. Inability of neonatal cells to induce an Mls-defined MLC was found not to be due to a low frequency of B lymphocytes or to the presence of suppressor cells, but due to the absence of the Mls-coded LADs on their surface. These data support the concept that the Mls-coded LADs are present on adult B lymphocytes and are specific markers of B-cell differentiation, which is preceded by membrane IgM and the δ homologue of human IgD, Ia, and the receptor for the third component of complement.     

Surface antigen changes occurring in short-term cultures of activated human T lymphocytes: analysis by flow cytometry

Surface antigens of activated and cultured human T cells were studied using peripheral blood lymphocytes activated with conditioned medium from phytohemagglutinin-activated leukocytes and maintained in liquid culture for 2 weeks with conditioned medium containing Interleukin 2. The ensuing cell population was tested for kinetic changes in cell size and for the expression of surface antigens by immunofluorescence staining with a panel of monoclonal antibodies and analysis by flow cytometry. Upon activation, the cell population progressively increased in size to large blasts, with the rapid appearance on all of the large dividing cells of the antigen recognized by OKT9, the transferrin receptor. Cells within the population continued to express the common peripheral T-cell antigens bound by OKT3 and UCHT1, and also the antigen bound by 3A1, but never the antigen bound by OKT6, a thymic cell marker. From the time of activation an increasing proportion of the T cells, up to 80%, expressed the antigen detected with OKIa and FMC4, which recognise nonpolymorphic Ia determinants. This sequence of events was followed by a general decrease in size of the cell population, a process accompanied by further phenotypic changes. The percentage of cells expressing Ia antigens decreased, but most striking was the rapid change in the OKT4:OKT8 ratio of cells within the population, from 60:40 to 40:60. Thereafter the proportions of OKT4⁺ to OKT8⁺ cells within the cultures remained relatively stable and it is suggested that these data provide evidence for a possible change in phenotype of cultured human T lymphoblasts, from OKT4 to OKT8.     

Potential identification of chemical carcinogens in a viral transformation system

Chemical carcinogens from several diverse chemical classes i.e.; aromatic amines, polycyclic hydrocarbons, nitrosamines, hormonal derivatives, metals and direct alkylating agents cause a 6.2–60.5-fold increase in the frequency of murine sarcoma virus (MSV)-induced transformation in a normal rat kidney (NRK) cell system. Exogenous metabolic activation with a rat liver S-9 homogenate is required for expression of this activity by procarcinogens. Non-carcinogenic analogs of these compounds fail to cause significant increases in the transformation frequency either with or without prior metabolic activation. Iododeoxyuridine, a mutagen also does not cause enhancement of transformation. This system may serve as the basis for a rapid and quantifiable means of identifying chemical carcinogens while introducing a new model for the understanding of the interactions between oncorna-viruses and chemical carcinogens.     

Replication and amplification of recombinant plasmid molecules as extra chromosomal elements in transformed mammalian cells

Mouse thymidine kinase negative (TK^?) L cells, F4-12B2TK^? Friend erythroleukemic cells and hamster BHKTK^? cells were transformed in the absence of carrier DNA with closed circular molecules of herpes simplex virus 1 TK DNA cloned in plasmid pAT153 (pTK-1). The physical status of donor DNA in the transformed cells was studied by Southern blot hybridization and spot hybridization techniques. Up to 65 copies of pTK-1 DNA molecules/cell were present in transformed cells grown under selective conditions. Whereas a steady increase in the number of pTK-1 copies/cell was found during the first few weeks of growth in selective medium, 2–8 months later copy numbers varied within the same cell line and their numbers rarely exceeded fifty copies/cell. Donor DNA sequences were found both in the Hirt precipitate and in the supernatant showing that some of the pTK-1 molecules existed in circular form. Many of the cell lines gave a Southern blot hybridization pattern indicative of pTK-1 sequences integrated into high molecular weight DNA as well as present in a circular configuration.     

Dental agenesis in the Dariusleut Hutterite Brethren: comparisons to selected Caucasoid population surveys

We report the results of a study of the prevalence of nonsyndromic dental agenesis among a sample of 208 individuals (105 females, 103 males) between the ages of 15 and 29 years from a North American religious and genetic isolate, the Dariusleut Hutterites of Western Canada. Direct examination of dental casts, oral examination reports, dental treatment records, and a limited number of dental radiographs reveals congenital absence and/or obvious morphometric reduction of at least one tooth (excluding third molars) in 98 subjects (55 females, 43 males), yielding a prevalence estimate of approximately 47%. This estimate is nearly four times those reported for nonisolate Caucasoid populations of European descent and substantially higher than the elevated prevalences observed in several other isolated populations. Although the prevalence of dental agenesis in the Dariusleut is indeed high, neither the incidence of bilateral agenesis (exhibited at least once in 58% of affected dentitions), number of affected teeth per person (mean, 2.4), morphologic tooth classes affected, or combinations of tooth classes affected ostensibly distinguish them from other populations with similar geographic origins. We conclude that the dental agenesis observed in this North American genetic isolate does not represent a private polymorphism or rare developmental variant. Consequently, the results of further study in these Dariusleut Brethren will be directly relevant to critically testing as yet unresolved hypotheses for the mode of gene action and the relative contributions of hereditary and environmental factors to the reduction of tooth numbers in human dentitions.     

A new variant of Autographa californica nuclear polyhedrosis virus

A variant of the baculovirus, Autographa californica nuclear polyhedrosis virus, has been isolated in Idaho during an epizootic disease in a field population of A. californica. Genotypic characterization indicates that the virus is distinct from variants previously characterized. Analysis of five clones, derived by plaque purification in cell culture, indicates relative homogeneity of the original virus isolate. Further exploration of the factors involved in natural genetic variability of baculoviruses is appropriate.