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To investigate the effect of lysosomal-associated protein transmembrane-4 Beta(Laptm4b) deletion on hematopoietic stem/progenitor cells (HSPCs) homeostasis in mice.The hematopoietic system specific Laptm4b-deficient mice were constructed. The number and proportion of HSPCs (LSK, LT, ST, MPP, etc) in Laptm4b-deficient mice were analyzed by flow cytometry. Single SLAM-HSC cell was sorted by flow sorter and cultured in vitro to measure the effect of Laptm4b deletion on the colony forming ability of hematopoietic stem cells (HSCs). The effect of Laptm4b-deficient on the reconstitution ability of HSCs in mice was detected by competitive transplantation experiment of SLAM-HSC cells.Laptm4b deficiency could moderately upregulate the proportion of T cells in the peripheral blood of the mice, but showed no significant effect on the proportion and number of HSPCs. Laptm4b deletion showed no effect on the reconstruction ability of HSCs after competitive transplantation, but it could inhibit the colony formation of HSCs in vitro.LAPTM4B may play a role in HSCs under the proliferation stress. Laptm4b-deficient in mice hematopoietic system showed no significant effect on the HSPCs homeostasis maintenance and reconstruction ability.Laptm4b缺失对小鼠造血干祖细胞稳态维持的影响.探讨溶酶体相关4次跨膜蛋白B（LAPTM4B）在小鼠造血干祖细胞稳态维持过程中的作用.构建造血系统特异的Laptm4b缺失小鼠；采用流式细胞术分析该模型小鼠的造血干祖细胞：LSK、LT、ST、MPP等细胞的数目和比例；通过流式分选仪分选造血干细胞SLAM-HSC，体外培养检测Laptm4b缺失对造血干细胞克隆形成能力的影响；通过造血干细胞SLAM-HSC竞争性骨髓移植实验分析Laptm4b缺失对小鼠造血干细胞造血重建能力的影响.Laptm4b基因缺失仅轻微上调稳态下小鼠外周血中的T细胞的比例，对稳态下小鼠造血干祖细胞的比例和数量无显著影响。Laptm4b基因的缺失不影响造血干细胞竞争性移植后的造血重建能力，但能够抑制体外培养下造血干细胞的克隆形成.Laptm4b缺失可能在造血干细胞体外扩增等快速增殖压力下发挥一定的抑制作用。在造血系统中特异敲除Laptm4b基因对小鼠造血干祖细胞的稳态维持及其造血重建能力无显著影响.

A 5423 bp fragment of LsNPV genome was sequenced, in which PDV-E66 gene and another four ORFs were found. The PDV-E66 gene of LsNPV was compared with the PDV-E66 gene of AcNPV, and a 51.9% nucleotide sequence homology and 38.8% amino acid sequence homology were found between the two genes. Two conserved late transcriptional motifs TAAG were found in LsNPV PDV-E66 gene, similar to those in AcNPV PDV-E66. The LsNPV PDV-E66 ORF is 204 base pairs shorter than the AcNPV PDV-E66 ORF at the 5′ end. This is agreement with the fact that the N-terminus of the AcNPV PDV-E66 mature protein is 69 amino acids interior to the N-terminus predicted by the AcNPV PDV-E66 ORF. The 5′ regulatory region of ORF1 contains early (CGTGC) and late (TAAG) transcriptional initiation motifs and ORF1 is predicted to encode a protein with 114 amino acid residues. The 5′ regulatory region of ORF2 which can encode a protein with 115 amino acid residues contains only an early transcriptional initiation motif. Compared with all the genes from AcNPV and other baculoviruses, ORF1 and ORF2 have no homologous genes. It is suggested that ORF1 and ORF2 may be two novel baculovirus genes. ORF3 (PDV-E66 gene), ORF5 and an incomplete ORF, ORF6-part, have homologous regions in the AcNPV genome. ORF3, ORF5, ORF6-part are linked together in LsNPV genome, but their homologous regions are separated by about 58 kb fragment in the AcNPV genome. This fact indicates that the organization of the above genes in LsNPV is different from that of AcNPV. ORF4 is included in ORF6-part and can encode a 48 amino acid residues polypeptide, but ORF4 and ORF6-part are located on different DNA strands.

A 1446 bp SalI fragment of LsNPV was sequenced by the silver staining method, and two large open reading frames (ORFs, ORF1 and ORF2) were found, both contain typical characteristics of the 5' regulatory elements of baculovirus early genes. ORF1 is 345 bp long with the capacity to encode a putative protein of 114 amino acid residues with MW about 13 kDa and was designated p13 gene, ORF2 comprises 248 bp from the 3' end of the fragment. In the untranslated region (UTR) of ORF1, a 33 bp mini cistron (ORF3), a core recognition sequence (CGTCG) for many bHLHzip transcription factors and a late promoter sequence TTAAG are present. In the UTR of ORF2, two host transcription factor binding elements (CACGTG and GATA motif) and two CGT motifs were found. Some regular leucine zipper-like structures, designated leucine trans-conformation structure and LVT repeat, were found near the N-terminus and the middle of p13 protein. The leucine trans-conformation structure that is near the N-terminus consists of 4 leucines and 7 other amino acids between every two leucines, and every leucine is located at a conformation shift point of the predicted secondary structure of the p13 protein. In LVT repeat, L-6aa-V-6aa-T-6aa is repeated once. The functions of those structures remain unclear, and the two ORFs, not found in the genome of Autographa californica nuclear polyhedrosis virus, are possibly two new genes.

A transfer vector was constructed in which the neomycin resistance (neo) gene was under the control of a copy of Autographa californica nuclear polyhedrosis virus (AcMNPV) IE1 gene promoter at the p10 locus. After cotransfection of Spodoptera frugiperda (Sf9) cells with the transfer vector and infectious AcMNPV DNA, the recombinant virus-containing neo gene was selected by serial passage of the mixed progenies from cotransfection. This was done at low MOI in the presence of G418 in growth medium and was followed by limited dilution. RNA dot hybridization showed that the neo gene was transcribed from immediately early phase to very late phase, in infected Sf9 cells. These results demonstrate that a new baculovirus vector system had been constructed in infected cells. Furthermore, a new method for selection of positive recombinant baculovirus had been developed.