Acute promyelocytic leukemia (APL) is characterized by a specific chromosome translocation involving RAR␣ and one of four fusion partners: PML, PLZF, NPM, and NuMA genes. To study the leukemogenic potential of the fusion genes in vivo, we generated transgenic mice with PLZF-RAR␣ and NPM-RAR␣. PLZF-RAR␣ transgenic animals developed chronic myeloid leukemia-like phenotypes at an early stage of life (within 3 months in five of six mice), whereas three NPM-RAR␣ transgenic mice showed a spectrum of phenotypes from typical APL to chronic myeloid leukemia relatively late in life (from 12 to 15 months). In contrast to bone marrow cells from PLZF-RAR␣ transgenic mice, those from NPM-RAR␣ transgenic mice could be induced to differentiate by all-trans-retinoic acid (ATRA). We also studied RARE binding properties and interactions between nuclear corepressor SMRT and various fusion proteins in response to ATRA. Dissociation of SMRT from different receptors was observed at ATRA concentrations of 0.01 M, 0.1 M, and 1.0 M for RAR␣-RXR␣, NPM-RAR␣, and PML-RAR␣, respectively, but not observed for PLZF-RAR␣ even in the presence of 10 M ATRA. We also determined the expression of the tissue factor gene in transgenic mice, which was detected only in bone marrow cells of mice expressing the fusion genes. These data clearly establish the leukemogenic role of PLZF-RAR␣ and NPM-RAR␣ and the importance of fusion receptor͞corepressor interactions in the pathogenesis as well as in determining different clinical phenotypes of APL.Acute promyelocytic leukemia (APL) is characterized in most patients by accumulation of promyelocytes containing a specific chromosomal translocation t(15;17) and a unique sensitivity to all-trans-retinoic acid (ATRA) treatment (1, 2). There have been three variant translocations reported, including t(11;17)(q23;q21), t(5;17)(q32;q21), and t(11;17)(q13;q21). These chromosomal translocations invariably involve the RAR␣ (retinoic acid receptor ␣) gene on chromosome 17, and the PML

The cellular prion protein (PrP C), a membrane glycoprotein anchored to the outer surface of neurons, lymphocytes and other cells, is associated directly with the pathogenesis of the transmissible spongiform encephalopathies (TSEs) occurring mainly in humans, cattle, sheep and goats. Although mice lacking PrP C develop and reproduce normally and are resistant to scrapie infection, large animals lacking PrP C , especially those species in which TSE occurs naturally, are currently not available. Here, five live PRNP +/" goats cloned by gene targeting are reported. Detailed RNA-transcription and protein-expression analysis of one PRNP +/" goat showed that one allele of the caprine PRNP gene had been disrupted functionally. No gross abnormal development or behaviour could be seen in these PRNP +/" goats up to at least 3 months of age. These heterozygous PRNP +/" goats are ready to be used in producing homozygous PRNP "/" goats in which no PrP C should be expressed.

This study was designed to produce cloned goats from cumulus cells. Cloning donor nuclei were from cumulus cells either freshly isolated or cultured in vitro. Enucleated oocytes were either injected with cumulus cell nuclei without piezo-driven manipulator (injection method) or fused with cumulus cells (fusion method). The survival rate of cloned embryos, obtained by injection, was higher than that derived from fusion (62.7 and 45.9%, respectively). Two cloned goats were derived by fusion with in vitro cultured cumulus cells without starvation, but died shortly after natural birth, from respiratory difficulties. Their birth weights (2.23 kg and 2.03 kg) were within the normal range (2.0-2.7 kg) and postmortem analysis revealed no morphological abnormalities. The third cloned goat, derived by injection of nuclei from freshly isolated cumulus cells, weighed 3.3 kg at birth, and was 37% overweight compared with the average weight of the same species. This goat is healthy and well as this paper is being prepared. Nested PCR-RFLP analysis confirmed that all the cloned goats were derived from the donor cells.

The neomycin-resistant gene (neo(r)) is probably the most commonly used selectable marker gene in gene targeting and gene transfection research. In this study, the neo(r) gene construct was introduced into in vitro cultured goat foetal fibroblast cells (IV-5), and the cells were selected with 900 microg/ml G418. The G418-resistant colonies were analysed by neo-specific PCR, karyotyping and anti-intermediate filament proteins antibody (anti-vimentin) staining. Cell cycle analysis of the neo(r) positive foetal fibroblast cell colony (IV-5.1) cultured in a variety of cell cycle-arresting medium indicated that 74.2% of cells cultured in serum-deprived medium for 3 days and 71.7% of cells grown to confluence were at G0/G1 stage of cell cycle, respectively, in comparison to 61.6% of cells in normal culture (cycling) medium. Nocodazole treatment for 17 hr in vitro culture could increase the number of cells at G2/M stage of cell cycle from 20.3% (in cycling medium) to 39.7%. In total, one early pregnancy was observed by B ultra-sound scanning in a surrogate transferred with cloned embryos from IV-5.1 cells at M stage (cells were cultured in nocodazole medium). Seven cloned goats, including two that miscarried at a late stage, were derived from the IV-5.1 cell clone cultured in starved medium (G0). Indeed, one surrogate receiving three blastocysts reconstituted from the starved donor cells, gave birth to three live cloned goats, all of which are healthy and doing well. PCR, Southern blot and G418 resistance in vitro of fibroblast cells from cloned goats confirmed that all cloned goats are positive for neo(r) transgene. This study demonstrates that a foreign gene, such as the neo-resistant gene, can be introduced into goat foetal fibroblast cells, and that the resulting transgenic cells are capable of being cloned to produce 100% transgenic animals.

Extracts of hairpencils ffom male cotton bollworm moth were analyzed by capillary gas chromatography, acid methanolysis, and GC-MS. Ten components have been identified as: 14 1 OH, 14 : Ac, 14 : COOH, 211-16 : OH, 16 t OH, 16 * Ac, 16 COOH, 18 1 OH, 18 t Ac, and 18 :COOH. Based on the statistics of titer of each chemical, the total amount of three saturated alcohols is over 75%. The amounts of the chemicals in the hairpencils are related to the age of males. There are no chemicals identified in the extracts from males less than 10 h after eclosion, then the quantity of compounds increased rapidly during 48 h after adult eclosion. After 5 days, the quantity decreased.

No abstract

Plant roots play critical roles in absorbing nutrients for the growth and development of plants as well as adapting different environments. Currently, there is not a satisfactory way to track dynamic information in the study of roots at the high temporal and spatial resolution. Herein, a simple microfluidic device with crossed microchannels was utilized for microscopic investigation of Arabidopsis thaliana roots in situ. Our experimental results showed that the microfluidic system combined with a microscope could be conveniently utilized for quantification of primary roots and root hairs with the change of micrometers within the time of minutes. With the same approach, influences of the stress of high salinity could also be investigated on different parts of the roots, including root cap, meristematic zone, elongation zone, mature zone, and root hairs, etc.More importantly, the growth of roots and root hairs could be quantified and compared in the solution of abscisic acid and indole-3-acetic acid, respectively. Our study suggested that the microfluidic system could become a powerful tool for the quantitative investigation of Arabidopsis thaliana roots.

AIM:To establish transgenic mice lineage harboring hepatitis B virus X gene and to provide an efficient animal model for studying the exact role of the HBx gene in the process of hepatocarcinogenesis.METHODS:The HBx transgenic mice were produced by microinjecting the construct with X gene of HBV (subtype adr) DNA fragment into fertilzed eggs derived from inbred C57BL/6 strain; transgenic mice were identified by using Nested PCR; expression and phenotype of HBx gene were analyzed in liver from transgenic mic at the age of 8 weeks by RT-PCR, pathologic examination and periodic acid-schiff staining (PAS), respectively.RESULTS:Five hundred and fourteen fertilized eggs of C57 BL/6 mice were microinjected with recombinant retroviral DNA fragment, and 368 survival eggs injected were transferred to the oviducts of 18 pseudopregnant recipient mice, 8 of them became pregnant and gave birth to 20 F1 offspring. Of 20 offsprings, four males and two females carried the hybrid gene (HBx gene). Four male mice were determined as founder, named X 1, X 5, X 9 and X 15. These founders were back crossed to set up F1 generations with other ibred C57BL/6 mice or transgenic littermates, respectively.Transmission of HBx gene in F1 offspring of X 1, X 5 and X 9 except in X 15 followed Mendelian rules. The expression of HBx mRNA was detected in liver of F1 offspring from the founder mice (X 1 and X 9), which showed vacuolation lesion and glycogen positive foci.CONCLUSION:Transgenic mice harboring HBx gene were preliminarily established.