The complete nucleotide sequence of a 16S ribosomal RNA gene from tobacco chloroplasts: Recombinant plasmid; sequence homology; stem and loop structure

The complete nucleotide sequence of a 16S ribosomal RNA gene from tobacco chloroplasts has been determined. This nucleotide sequence has 96% homology with that of maize chloroplast 16S rRNA gene and 74% homology with that of Escherichia coli16S gene.The 3′ terminal region of this gene contains the sequence ACCTCC which is complementary to sequences found at the 5′ termini of prokaryotic mRNAs.The large stem and loop structure can be constructed from the sequences surrounding the 5′ and 3′ ends of the 16S gene. These observations demonstrate the prokaryotic nature of chloroplast 16S rRNA.     

Synthesis of RNA polymerase in Escherichia coli B-r growing at different rates

Polyacrylamide gel electrophoresis of unfractionated sodium dodecyl sulfate lysates of Escherichia coli$\text{B}\text{r}$ has been used to investigate the synthesis of β and β′ subunits of RNA polymerase as a function of bacterial growth rate. In succinate (μ = 0.67 doublings/h), glucose (μ = 1.36 doublings/h) and glucose/amino acids (μ = 2.10 doublings/h) supplemented media, the fraction of [¹⁴C]leucine-labeled β and β′ protein/total protein was found to be 1.05, 1.31 and 1.56%, respectively. Comparison of these values with recent estimates from this laboratory of the differential rate of synthesis of functioning RNA polymerase suggests an excess of total over functioning RNA polymerase. The significance of these data in reference to the regulation of RNA polymerase synthesis is discussed.     

Nucleotide sequence of the chromosomal gene for human fibroblast (beta 1) interferon and of the flanking regions

The nucleotide sequence of the human fibroblast (beta 1) interferon chromosomal gene and its flanking regions was determined. These results confirm the absence of intervening sequences in the gene. The presence of some sequences in the upstream flanking region homologous to similar features for other eukaryotic genes was revealed: these include not only the TATAAAT sequence and the consensus sequence (reported by Benoist et al., 1980) but also two additional motifs, one of which is so far present only in inducible genes. Furthermore, a striking similarity between the upstream flanking regions of the human beta 1 and alpha 1 interferon genes is observed.     

Effect of adenosine deaminase inhibitors on Fcμ receptor expression in human T-cell cultures

Incubation of human peripheral blood T-lymphocyte cultures with 100 μM EHNA or 1 μM pentostatin results in the delayed appearance of Fcμ, receptor-bearing cells. The percentage of Tμ-positive cells approaches control levels by 24–30 hr despite the lack of detectable adenosine deaminase activity. Tγ to Tμ transition was more effectively inhibited than To to Tμ in studies with the individual subpopulations. The inhibitory effect of pentostatin or EHNA on the appearance of Fcμ receptor-positive cells was reversible with exogenous undine or 2′-deoxycytidine. These results indicate that pyrimidine deprivation affects the appearance of Fcμ receptors in T-cell cultures, and that despite the continued inhibition of ADA the effect on Fcμ is reversible by 24 hr without the addition of exogenous pyrimidines.     

Optical activity of single-stranded polydeoxyadenylic and polyriboadenylic acids; dependence of adenine chromophore cotton effects on polymer conformation

Circular dichroism (CD) curves are reported for poly dA, (pdA)₆, (pdA)₂, poly A, ApAp, ApA, AMP, dApA, pdApA, A-2′-O-methyl pA, and A-2′-O-methyl pAp. Analysis of these curves indicated the presence of single CD bands at 228–230 mμ and at 278–280 mμ in oligomers longer than dinucleotides. In the case of dinucleotides and mononucleotides (from the literature, in addition to those studied here), the 230 mμ CD of band appears but the 280 mμ CD band does not. We assign the 230 mμ band to a very weak π–π* transition at this wavelength. From theoretical considerations, we show that the 280 mμ band is not an exciton component of the strong π–π* transition at 260 mμ in adenine. We conclude that the 280 mμ CD band must be assigned to a distinct absorption, not previously reported, which we suggest arises from an n–π* transition. The fact that the n–π* CD band at 280 mμ is not seen in mononucleotides or dinucleotides is ascribed to solvation of the adenine ring by water, which shifts the band to shorter wavelengths. Therefore, only interior residues of oligomers have the 280 mμ band, and the optical activity of a polymer cannot be computed from that of a dinucleotide, by using a nearest-neighbor approximation. The existence of this end effect hag been tested, by taking it into account in computing the rotational strengths of the 278 mμ n–π* transition for several oligomers; it is pointed out that a more sensitive test of this end effect would require CD data for the oligo dA series of 3 to 5 residues. We speculate about the structural and optical differences between poly dA and poly A, and point out the need for a theoretical treatment of n–π* Cotton effects in polynucleotides.     

耐辐射奇球菌超氧化物歧化酶基因的克隆与序列分析

By using a 453 bp length gene fragment of superoxide dismutase(SOD)as a probe,which was firstly amplified from Deinococcus radiodurans genomic DNA by PCR with degenerate oligonucleotide primers corresponding to the conservative regions of known SODs,a putative SOD gene was identified from the database of D.radiodurans whole genome.Its 636 bp length open reading frame and 5′ and 3′ flanking sequence was determined.The conventional E.coli ribosomal and RNA polymerase binding sites were found upstream from SOD encoding region and an inverted repeat sequence downstream of the termination codon.The deduced 211 amino acid sequence of the structural gene showed a high similarity to other manganese and iron containing SODs in normally conserve regions.     

Nucleotide sequence of the dihydrofolate-reductase gene borne by the plasmid R67 and conferring methotrexate resistance

The complete nucleotide sequence of the methotrexate-resistant dihydrofolate reductase (DHFR) gene borne by the plasmid R67 was determined. The gene is 234 bp long and codes for 78 amino acids. The polypeptide deduced from the DNA sequence is in perfect agreement with the previously published amino acid sequence. Comparison of the nucleotide sequence with the one determined for the R388-encoded DHFR indicates that 75% of the nucleotides are conserved in the two genes. The 3′ end of the R67 gene can be modified without altering significantly the activity of the enzyme.     

我国两株登革2型病毒5′和3′端非编码区序列测定及二级结构分析

 为测定我国两株临床症状、乳鼠神经毒力不同的登革 2型病毒流行株 5′和 3′端非编码区序列 (untranslated region,UTR) ,分析二级结构差异与毒力变化的关系 ,分别从 D2 - 0 4、D2 - 44株感染的 C6/ 36细胞及鼠脑中提取总 RNA.以该 RNA为模板 ,利用 RACE法 ,分别扩增了 D2 - 0 4、D2 -44株的 5′和 3′末端 c DNA片段 .将其分别与 p GEM- T载体连接得到重组质粒 ,测定上述 c DNA插入片段的序列 .用 RNAdraw软件预测 D2 - 0 4、D2 - 44株 5′和 3′端非编码区的二级结构 .D2 - 0 4、D2 -44株 5′端和 3′端非编码区分别有 96和 454个核苷酸 .其中 5′非编码区 59位 C(D2 - 0 4 )→T(D2 -44 ) ,使 D2 - 44二级结构稳定性下降 ;3′端非编码区有 1 5个核苷酸不同 ,其中 T(355)→ A,T(32 6)→ G引起了所在位置二级结构自由能变化 ,且分别位于两个保守序列区 (conserved sequence,CS)CS1、CS2 A.这些位点变化可能与毒力有关 .     

茶树肌动蛋白基因（CsActin1）全长cDNA克隆与生物信息学分析

以茶树（Camellia sinensis）萌动芽为材料,根据茶树萌动芽芽抑制消减杂交文库中分离得到的肌动蛋白（actin）基因的5′-片段设计引物,利用3′-RACE技术克隆了其cDNA全长序列,该基因cDNA全长1 470 bp,命名为CsActin1（GenBank登录号HQ235647）。序列分析表明,CsActin1开放阅读框长1 134 bp,编码377个氨基酸,5′非编码区100 bp,3′非编码区236 bp。推测的蛋白质分子量为41.70 kD,等电点约为5.31,具有肌动蛋白家族的特征信号序列（YVGDEAQs.KRG和WIAKaEYDE）和肌动蛋白相关蛋白的特征信号序列（LLTEApLNPkaNR）。CsActin1与GenBank中注册的其它植物肌动蛋白核苷酸序列的相似性在80%以上,氨基酸序列相似性在95%以上。与其它植物肌动蛋白的进化树分析结果表明,茶树肌动蛋白与杨树的两个肌动蛋白间的亲缘关系最为密切。并对推导的蛋白结构进行了分析。     

血红密孔菌(Pycnoporussanguineus)漆酶基因的克隆与序列分析

为克隆血红密孔菌 (Pycnoporussanguineus)漆酶基因 ,根据真菌漆酶氨基酸序列保守区设计了 1对简并引物 .以血红密孔菌基因组DNA为模板 ,PCR扩增出长 12 2 7bp的漆酶基因片段 .以此序列为基础 ,通过 5′及 3′RACE技术克隆出漆酶全长cDNA序列 ,序列长为 190 2bp ,其 5′端和 3′端非编码区长分别为 5 1bp和 2 97bp ,开放阅读框长 15 5 4bp ,编码 5 18个氨基酸的蛋白 .该蛋白具有 4个铜离子结合区域 ,预测其相对分子量为 5 6 313 2 ,等电点为 5 5 9,其氨基酸序列与Pycnoporuscinnabarinus漆酶 (lcc3 2 )的同源性最高 ,为 96 % .以该cDNA编码区的两端序列为引物 ,PCR扩增得到漆酶的长度为 2 15 4bp的全长DNA序列 ,序列中包括 10个内含子序列 ,长为 5 2～ 70bp     

Bovine Epizootic Fever

It was found that the ruins of Bovine Epizootic Fever appeared to be RNA as its replication was not inhibited by 5-iodo-2′-deoxyuridine. It had a buoyant density of 1.196, was not filterable through membrane filter (Millipore Filter Corp. Mass.) of 100 mμ pore size, and was sensitive to ether, chloroform and deoxycholate, was inactivated by trypsin and ultraviolet irradiation, precipitated by protamine sulfate, readily inactivated at pH 3.0, fairly labile at 56 C but readily preserved at —80 C, not stabilized at 50 C by I M MgCl₂, and resisted repeated freeze-thawing. The virus appears not to require a DNA-depcndent RNA synthesis in the host cell for its replication, as chromomycin A3 did not inhibit its replication. Sucrose density gradient centrifugation was unsuitable for purification because of a very poor recovery of infectivity. CsCl equilibrium density gradient centrifugation was successfully used for this purpose, and electron microscopy of the resulting fractions by phosphotungstic negative staining technique revealed virus-like bullet-shape particles, about 140 mμ in length and 80 mμ across, in the fraction of peak infectivity titer. The particle is probably the virion of the virus. These findings suggest relation of the virus to Rhabdoviruses [6], but further studies are necessary. The physicochemical properties of the virus provide additional evidence for similarity of the virus to bovine ephemeral fever virus, and emphasizes the desirability of further detailed comparative studies to decide whether they are one and the same or merely very closely related.     

Ibaraki Virus,an Agent of Epizootic Disease of Cattle Resembling Bluetongue

Replication of Ibaraki virus was not inhibited by 5-iodo-2′-deoxyuridine, indicating that the virus is an RNA virus. The virus was resistant to ether, chloroform and deoxycholate, sensitive to trypsin, very labile at acidic pH but stable at pH 6.4 or higher, and was resistant to repeated freezing and thawing. The virus was readily inactivated at 56 C or higher, was fairly stable at 37 C, and very stable at 4 C, while it rapidly lost infectivity when stored frozen at —20 C. The virus was readily sedimented by centrifugation at 40 000Xg for 60 min. It readily passed through membrane filters of 200 mμ pore size, passed through 100 μfilters but only with some titer loss and did not through 50 mμ filters. In these tests, the bluetongue virus used as a control behaved in the same manner as Ibaraki virus. These findings provide additional evidence for the similarity of Ibaraki virus to bluetongue virus which had been previously demonstrated on the basis of seasonal incidence, symptomatology and pathology of the diseases caused by these viruses and the behavior of the viruses in cell cultures, embryonated eggs and laboratory animals. The present study, however, provided no evidence for any serological relation between these two viruses. More Information is needed to reach a final decision on the classification of Ibaraki virus, particularly regarding the morphology of the virion, the doublestrandedness of the viral RNA and other basic features.     

Nucleotide sequence and exact localization of the neomycin phosphotransferase gene from transposon Tn5

The nucleotide sequence of 1200 bp from the unique region of transposon Tn5 containing the neomycin phosphotransferase gene (neo) was determined, and the location of the neo gene was identified by deletion mutants in a translational reading frame of 792 bp. The derived gene product, an aminoglycoside 3′-phosphotransferase (APH) II, consists of 264 amino acid residues and has a calculated M_r of 29053. Its amino acid sequence shows sequence homologies to the APH type I enzyme coded for by transposon Tn903 (Oka et al., 1981).     

PHOSPHATASE ACTIVITY AND CELLULAR DIFFERENTIATION IN PHLEUM ROOT MERISTEM

Using 9 different organic phosphate substrates as alternatives in a standardized 5′-nucleotidase histochemical test system, enzyme activity patterns were recorded for timothy grass root epidermis. At least 4 different phosphatases were distinguished on the bases of substrate specificity, reaction rate, tissue distribution, and response to inhibitors. Except with adenosine-3′-monophosphate, all activities were restricted to the 300-mμ-long root tip meristem. These enzyme activities were associated with the earliest phases of differentiation of the epidermal hair and hairless cell initials. The distribution of activities was not associated with the same cell type in each part of the meristem. Little activity was found with most substrates in the undifferentiated cells of the 0-100μ zone; alternating active hairless and inactive hair cell initials predominated in the 100-200μ segment; and active hair–inactive hairless sister cells formed the principal pattern in the 200-300μ segment of the meristem. The data showed that a particular enzyme activity was associated with a specific cell type only in relation to that cell's position along the differentiation gradient of the entire tissue. But, within a meristem segment, a specific cell type might act differently from its neighbors, depending on its mitotic capacity. This complex of physiological dependence and independence of a cell type on tissue ontogeny was cited as a characteristic of the phenomenon of cellular differentiation superimposed on tissue differentiation gradients.     

Two forms of loops generate the chromatin conformation of the immunoglobulin heavy-chain gene locus

The immunoglobulin heavy-chain (IgH) gene locus undergoes radial repositioning within the nucleus and locus contraction in preparation for gene recombination. We demonstrate that IgH locus conformation involves two levels of chromosomal compaction. At the first level, the locus folds into several multilooped domains. One such domain at the 3' end of the locus requires an enhancer, Eμ; two other domains at the 5' end are Eμ independent. At the second level, these domains are brought into spatial proximity by Eμ-dependent interactions with specific sites within the V(H) region. Eμ is also required for radial repositioning of IgH alleles, indicating its essential role in large-scale chromosomal movements in developing lymphocytes. Our observations provide a comprehensive view of the conformation of IgH alleles in pro-B cells and the mechanisms by which it is established.     

Transcripts coding for variant surface glycoproteins of Trypanosoma brucei have a short,identical exon at their 5′ end

The nucleotide sequence of the 5′ end of the mRNA for variant surface glycoprotein (VSG) 117 has been determined and compared with the sequence of the unexpressed basic copy (BC) of the VSG 117 gene. This shows the existence of an exon 35 nucleotides long at the 5′ end of the mRNA. The evidence suggests that this ‘mini-exon’ is derived from the expression site into which the VSG 117 BC is transposed during activation. The nucleotide sequence of this mini-exon is indistinguishable from that recently found for a different VSG, 118 (Van der Ploeg et al., Nucl. Acids Res. 10 (1982) 3591–3604). Analysis of the 5′ end of the mRNA for another VSG (221) whose gene is thought to be activated by a different mechanism to that of VSGs 117 and 118 indicates that the 5′- most 35 nucleotides of the VSG 221 mRNA are identical to the 117/118 mini-exon sequence. The implications of these results for the mechanism of VSG gene expression are discussed.     

ggpps 5′-侧翼序列的克隆及其启动子功能验证

目的：研究栊牛儿基栊牛儿基焦磷酸合成酶（geranylgeranyl diphosphate synthase,GGPPs）基因启动子的活性;方法：从曼地亚红豆杉细胞中克隆ggpps基因5′-侧翼序列,并将该侧翼序列代替pBI121质粒上的CaMV35S启动子,以Gus基因作为报告基因构建植物表达载体,并进一步导入农杆菌LBA4404中获得阳性转化子,然后用叶盘转化法验证该侧翼序列的启动子活性;结果：本研究从曼地亚红豆杉细胞中成功克隆了ggpps基因的5′-侧翼序列,并且验证了该侧翼序列具有启动子活性;结论：ggpps基因的5′-侧翼序列的测序结果表明本实验成功克隆了该侧翼序列,启动子功能验证结果表明ggpps 5′-侧翼序列具有启动子活性,这些结果为进一步的通过缺失法进行ggpps基因启动子功能研究奠定了基础。     

原核增强子样序列3A的克隆及其结构与功能的研究

 从大肠杆菌 C60 0株染色体基因组中筛选到一个原核增强子样序列 3A,其正、反向增强活性分别能提高 β-半乳糖苷酶活性 7.1 1和 2 .93倍 .采用体内转录 ,RNA斑点印迹杂交的方法 ,证明3A序列对于基因表达的调控发生在转录水平上 . 3A功能区的研究表明 ,3A增强活性主要位于距其正向克隆 5′端 (增强活性较强的方向称为正向 ,反之为反向 ) 30 0～ 540 bp,长约 2 4 0 bp的区段内 ,并证明 3A片段增强活性至少由两个增强活性位点组成