Interactions between HMG proteins and the core sequence of DNaseI hypersensitive site 2 in the locus control region (LCR) of the human β-Mike globin gene cluster

HMG proteins are abundant chromosomal non-histone proteins. It has been suggested that the HMG proteins may play an important role in the structure and function of chromatin. In the present study, the binding of HMG proteins (HMG1/2 and HMG14/17) to the core DNA sequence of DNasel hypersensitive site 2 (HS2core DNA sequence, -10681-10970 bp) in the locus control region (LCR) of the human β-like globin gene cluster has been examined by using both the in vitro nucleosome reconstitution and the gel mobility shift assays. Here we show that HMG1/2 can bind to the naked HS2core DNA sequence, however, HMG 14/17 cannot. Using the in vitro nucleosome reconstitution we demonstrate that HMG14/17 can bind to the HS2core DNA sequence which is assembled into nucleosomes with the core histone octamer transferred from chicken erythrocytes. In contrast, HMG 1/2 cannot bind to the nucleosomes reconstituted in vitro with the HS2core DNA sequence. These results indicate that the binding patterns between HMG proteins and t     

扬子鳄的CaSox4基因的分子克隆和进化分析

The completely identical HMG-box motif of CaSox4 gene from both male and female genomic DNA of the Chinese alligator (alligator sinensis) was cloned and sequenced by degenerate primer PCR.Compared with the human and mouse SRY,CaSox4 revealed 51% and 57% nucleotide homology respectively and 49% and 55% amino acid identity respectively,CaSox4 belongs to subgroup C of the Sox gene family.The GC content is 86% in the HMG-box region of the CaSox4 gene,Blast analysis showed that the CaSox4 gene shares 100^ amino acid identity with human Sox4.bird SoxLF4,turtle Sra4 and lizard CvSox4 genes.Casox4 may be orthologous with the human SOx4 gene.This indicates that CaSox4 gene shows the remarkable evolutionary conservation during the evolution of Alligator Sinensis,The extensive sequence conservation of the Sox4 gene between reptiles,mammals and birds suggests major functional constraints[Acta Zoologica Sinica 49(3):404-407,2003].     

A novel gene——samfR involved in early stage of Streptomyces ansochromogenes differentiation

A 4.6 kb DNA fragment was cloned from the DNA library of Streptomyces ansochromogenes using a partial DNA fragment located in the downstream of promoter-P_(TH4) as probe. The experiments revealed that this DNA fragment consists of saw D gene and a 1.4 kb Pvu Ⅱ fragment which can accelerate mycelium formation of S. ansochromogerms. The nucleofide sequence of 1.4 kb DNA fragment was determined and analysed; the result indicated that the fragment contains one complete open reading frame (ORF) which encodes a protein with 213 amino acids, and this gene was desiguated as samfR. The deduced protein has 36% amino acid identities and 52% amino acid similarities in comparison with that encoded by hppR gene, which is involved in the regulation of catabolism for 3-(3-hydroxyphenyl) propionate (3HPP) in Rhodococcus globerulus. The function of samfR gene was studied using strategy of gene disruption, and the resulting samfR mutant failed to form aerial hyphae and spores, its development and differentiation stopped     

Structure and function ofsawB, a gene involved in differentiation ofStreptomyces ansochromogenes

A partial DNA library of Streptomyces ansochromogenes 7100 was constructed by using plasmid plJ702 as vector and white mutant W19 as recipient. About 3 000 clones were obtained, two of which gave rise to the grey phenotype as wild type 7100. The plasmids were isolated from two transformants. The result indicated that the 5.2 kb and 5.8 kb DNA fragments were inserted into plJ702. The resulting recombinant plasmids were designated as pNL-1 and pNL-2 respectively. The 1.25 kb Pstl l-Apa l DNA fragment from pNL-1 was recognized as its complementarity to W19 strain. The nucleotide sequence of the 3.0 kb Pst I DNA fragment including 1.25 kb was determined and analyzed. The result indicated that this DNA fragment contains one complete open reading frame (ORF1) which encodes a protein with 295 amino acid residues, and this gene was designated as sawB. The deduced protein has 81% amino acid identities in comparison with that encoded by whiH in Streptomyces coelicolor. The function of sawB gene was studied by usi     

RNA-binding domain of the key structural protein P7 for the Rice dwarf virus particle assembly

The Rice dwarf virus (RDV) P7 structural protein is the key protein in the RDV particle assembly. The P7 protein was digested partially or completely by Staphylococcus aureus V8 protease and/or Pseudomonasfragi Asp-N protease. The molecular mass and the N-terminal amino acid sequence of the polypeptide fragments of the P7 protein were determined by SDS-PAGE and the Edman degradation method, respectively. Then the polypeptides were located in the deduced amino acid sequence of the RDV P7 protein based on the nucleotide sequence information, with the knowledge of the specific cleavage sites of the Staphylococcus aureus V8 and Pseudomonasfragi Asp-N protease, and the two RNA-binding domains in the P7 protein were identified. Domain 1 was located in the residue 128-249 containing 122 amino acids and domain 2 was located in the residue 325-355 containing 31 amino acids. Thus, these two domains may play an important role in the virus particle assembly by contributing to the packaging of viral dsRNAs inside the particles. The two domains may be novel RNA-binding domains, because no amino acid sequences highly similar to the conservative sequences of known dsRNA-binding domains reported so far. The similarity between the motif of domain 1 and the motif of the DNA-binding protein suggests that the DNA-binding activity of the RDV P7 protein may be due to this sequence. The similarity between the motif of domain 1 and the motif of the RNA polymerase domain suggests that the P7 protein may also play a role in RNA synthesis, besides its function in the assembly and subsequent packaging of viral dsRNA into core particles.     

Antisense-Mediated Depletion of Tomato Chloroplast Omega-3 Fatty Acid Desaturase Enhances Thermal Tolerance

A chloroplast-localized tomato （Lycopersicon esculentum Mill.） ω-3 fatty acid desaturase gene （LeFADT） was isolated and characterized with regard to its sequence, response to various temperatures, and function in antisense transgenic tomato plants. The deduced amino acid sequence had four histidine-rich regions, of which three regions were highly conserved throughout the whole ω-3 fatty acid desaturasegene family. Southern blotting analysis showed that LeFAD7was encoded by a single copy gene and had two homologous genes in the tomato genome. Northern blot showed that LeFAD7 was expressed in all organs and was especially abundant in leaf tissue. Meanwhile, expression of LeFAD7 was induced by chilling stress （4 ℃）, but was inhibited by high temperature （45 ℃）, in leaves. Transgenic tomato plants were produced by integration of the antisense LeFAD7DNA under the control of a CaMV35S promoter into the genome. Antisense transgenic plants with lower 18 ： 3 content could maintain a higher maximal photochemical efficiency （Fv/Fm） and O2 evolution rate than wild-type plants. These results suggested that silence of the LeFAD7 gene alleviated high-temperature stress. There was also a correlation between the low content of 18 ： 3 resulting from silence of the LeFAD7 gene and tolerance to high-temperature stress.     

Human augmenter of liver regeneration: molecular cloning, biological activity and roles in liver regeneration

The complete amino acid sequence of human augmenter of liver regeneration (hALR) was reported by deduction from nucleotide sequence of its complementary DNA . The cDNA for hALR was isolated by screening a human fetal liver cDNA library and the sequencing of this insert revealed an open reading frame encoding a protein with 125aa and highly homologous (87% ) with rat ALR encoding sequence. The recombinant hALR expressed from its cDNA in transient expression experiments in cos-7 cells could stimulate DNA synthesis of HTC hepatoma cell in the dose-dependent and heat-resistant way. Northern blot analysis with rat ALR cDNA as probe confirmed that ALR mRNA was expressed in the normal rat liver at low level and that dramatically increased in the regenerating liver after partial hepatectomied rat. This size of hALR mRNA is 1.4 kb long and expressed in human fetal liver, kidney and testis. These findings indicated that liver itself may be the resource of ALR and suggested that ALR seems to be an im-portant parac     

番茄烟粉虱传双生病毒PCR检测

From the conserved regions of the reported nucleotide sequences of whitefly-transmitted geminiviruses (WTGV), a pair of degenerate primers was designed to anneal to the conserved sequence.The tomato samples infected geminivirus-like from Guangdong were detected by PCR. The results showed that a 356bp specific fragment was amplified from the samples. The specific fragment was cloned and sequenced, and the sequence was compared with all nucleotide sequences in GenBank by Blast of NCBI. The result showed that the fragment belonged to Geminiviridae DNA. So the degenerate primers may be used to detect the WTGV from tomato in Guangdong. Moreover, both of the homology of the fragment between WTGV from tomato in Guangdong and the reported WTGV in the world and WTGV from tomato in Guangxi were under 82%. These results implied that the WTGV from tomato in Guangdong differed from the above-mentioned WTGV.     

Visualization of interaction between ribosome-inactivating proteins and supercoiled DNA with an atomic force microscope

The interaction between ribosome-inactivating proteins (RIPs) and supercoiled DNA was observed with an atomic force microscope (AFM). It was found that RIPs can bind to both supercoiled DNA and the unwound double stranded loop region in supercoiled DNA. The RIPs hound to the supercoils can induce the conformational change of supercoiled DNA. Furthermore, the supercoiled DNA was relaxed and cleaved into nick or linear form by RIPs. It indicated that RIP seemed to be a supercoil-dependent DNA binding protein and exhibited the activity of su-percoil-dependent DNA endonuclease.