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Stem cell factor (stem cell factor, SCF), a proto-oncogene c-kit ligand, role in hematopoiesis earlier hematopoietic growth factors, their gene positioning in the human chromosome 12 q 22 to 24 < / sup> District. SCF in vivo exist in two ways: the membrane-bound and soluble, soluble human SCF molecule, composed of 1 to 164 amino acids of the extracellular region, sometimes formed by non-covalent bond dimer exists in vivo, is a multifunctional cytokines acting on the multi-lineage hematopoietic cell. Since the SCF was discovered in 1990, its functions continue to be discovered, was clarified. SCF can be synergistically other cytokines, such as the EPO, of GM-CSF of G-CSF, IL-2, IL-3, IL-11 and TPO, SCF has a significant synergistic effect. The clinical application of SCF treatment of anemia, improve put the number of white blood cells after chemotherapy, in vitro hematopoietic gene therapy is a promising cytokine drugs. Has been identified in E.coli, unglycosylated soluble human SCF, with natural soluble human SCF has the same biological effects. Recombinant human stem cell factor in E.coli difficult to achieve high expression, from 1992 began to study expression of hSCF in E.coli, in order to obtain high-level expression, we have tried many methods, although did not achieve satisfactory results, but accumulated a lot of experience. Temperature sensitive expression vector of the present study pBV-220 as a template, and by means of computer analysis software to analyze the results obtained by the original work, and looking for the efficient expression of the factors to influence rhSCF in E.coli. pBV-220 expression vector, the the domestic scientists built prokaryotic expression vector, currently in the country are widely used in the production of recombinant drugs. Generally considered the influence of exogenous genes in the vector expression efficiency is primarily three factors: the SD sequence length, gene start code ATG and terminate at the code TAA RNA secondary structure and codon bias. SD sequence length can not afford a critical role in expression hSCF SD sequence length change not improve the expression efficiency; natural hSCF both ends of the RNA secondary structure in line with the high expression discriminant function, the test was highly expressed in analysis: usually purposeful transformation of exogenous gene vector at RNA secondary structure can be highly expressed modify hSCF both ends of the RNA secondary structure, did not achieve the high expression, suggesting that there may be another reason for existence. We believe that is due to the codon bias the affected gene hSCF efficient expression in E.coli. Soluble hSCF protein of 164 amino acids, including 57 amino acids codon the expression coefficient is lower than 0.1, 34.8% of the total number of amino acids. Since the codon the coding hSCF genes not E.coli preferred index of amino acids, and low expression group exists, causing the gene in E.coli difficult to achieve high _ the first. Military Medical University, Bo ten degrees of the righteous, \and efficient expression of the gene of hSCF a transformation, modify encoding hSCF amino acid part codon so the coding hSCF expression of each amino acid the 8_ index is not less than 0 * Segmented synthesis transformation gene, PCR disposable charge sake of gene splicing, clump Z-g Yinke Long Diao claw 220 carrier. sequencing results showed that the inserted gene sequence with the original design consistent expression after Shan Shen F *% share of the total bacterial protein induced g the genetically modified Lee before the expression efficiency of 10% I ratio, bamboo ￥ nearly 2-fold increased Western seven * identification, protein expression for hSCF monoclonal antibody identification f_ do a small amount of fermentation, preliminary purification of rhSCF protein purity greater than 80%, to promote Mo7e cells - proliferation activity. 'since 1998, erythroid differentiation, and constantly progress, erythroid differentiation mechanism is g was gradually clarified. SCF and its ligand Ren seven erythroid differentiation, it is not just co-erythrocyte students j into Su * rythrpoietin, the EPO and involved, and play an important role in recent studies of hair _ now, S * F * PO erythroid differentiation play a very important role, it is possible to suppress erythropoiesis twist _ cells apoptosis, promote erythroid dry, proliferation and differentiation of progenitor cells and erythroid progenitor cell growth state, the necessary conditions of differentiation-_ But S * F cells of normal development, whether inhibiting apoptosis, promoting the role of the increase, it? 8 ** L rat leukemia hematopoietic leaves., ** F has two of Lee threatened with the biological effects of the mountain] also has both stimulate normal hematopoiesis, but also can inhibit the proliferation of leukemic cells in the body SCF enough whether _-with two different biological effects? erythroleukemia cell line K562 cells for research template, try to explore the SCF small kit in erythroid l / progenitor cell proliferation and differentiation. has a large number of reported, hEpO HCmill and RA can lure - guide K562 cells to erythroid differentiation induced K562 cells, hemoglobin increased expression erythroid _ transcription factor activation, erythroid progenitor cells some of the characteristics of this study of rhEPO Flemin _ and RA K562 cells induced to erythroid differentiation, the prosecution?
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