| [Background]For Human Genome Project (HGP), one of important tasks is try toseparate the genes specifically expressed in a certain cell. It helps to not only predict the gene's location in chromosome but also analysis the gene structure and function in genome. More important, it can provide the foundation to elucidate the function of cells.All-trans retinoic acid (ATRA) is the earliest and most extensively used differentiation-inducing agent of malignant tumor in clinic up to now. But to attain a last anti-tumor effect, it must be used to combine with chemotherapeutic agents. At present, there still exit the dispute to the possibility of drug-resistance induced by ATRA. Though some researchers did observe the phenomena of drug-resistance of malignant tumor cells induced by ATRA, but due to the possible influence to cell divide and cell cycle by ATRA, the research is very poor. At present, most of the researches were only confined to the study of the mechanism and the reverse of the differentiation resistance by ATRA itself, while the research of multidrug-resistance to all kinds of chemotherapeutic agents reduced by ATRA was neglected. In a previously study, we ever observed the multidrug-resistance in hepatoblastoma cell induced by ATRA, and found the mechanism might not only relate to a certain multidrug-resistaht gene. [Purpose]For the sake of excluding the interferences, this experiment try to avoid the effect of cell divide, the cell cycle and the speciality of tumor cells, which were the factors probably exist in experiment. Then the human acute myeloid leukemia cell line (HL-60), which is the most common used as a model in the study of differentiation, was used as research object .A long-term, intermittent, inefficient ATRA was used to induce the tumorcell's resistance and try to establish a multidrug-resistance cell line, leaving out the interference of cell divide, the cell cycle and the speciality of tumor cells, then we try to make clear if ATRA can cause the multidrug resistance of malignant tumor cell. If it can, we would try to screen and separate the related gene of multidrug resistance in tumor cells, and provide an important information on how to overcome the multidrug resistance by ATRA in clinic. [Method]1 A long-term, intermittent, inefficient ATRA was used to inducetumor cell's resistance and try to establish a multidrug-resistance cell line (HL-60/ ATRA).2 The suppression subtractive hybridization library of HL-60/ATRA was established by using the HL-60 cells' cDNA as tester, while the HL-60/ATRA cDNA as driver.3 A high though screening technique, gene chip, was used to screen the differentially expressed genes in the established subtractive library.4 The differentially expressed gene was sequenced and analyzed by bioinformatics.[Results]1 The HL-60/ATRA could keep the undifferentiated states and its intrinsic proliferation in the appearance, biochemical metabolism and cell function at a high acceptable concentration (10-7M ATRA), but it became multidrug resistance to a variety of chemotherapeutic agents such as vincristine (VCR), adriamycin (ADM), mitomycin (MMC), daunorubicin (DNR), cyclophosphamide (CTX), et.al, the resistant factor varied from 3 to 10 times.2 A subtractive library was established by the suppression subtractive hybridization using the cDNA of HL-60 as "driver", while the cDNA of HL-60/ATRA as "tester" to screen the differentially expressed gene in HL-60 cells. The products of second subtractive hybridization were cloned into the vector of pUC57/T to create a subtractive library.510 Clones were selected from it and were identified by PCR. 433 clones of these clones were approved to contain a specific enhancive band with 200700 bps. The positive ratio was amount to 84.8%.3 To perform a high throughput analysis of cDNA clones with the intent of identifying genes expressed in association with the MDR phenotype in HL-60/ATRA cell lines, the cDNA inserts of 433 clones were amplified with...
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