In Bacillus subtilis, selenocysteine tRNA has not been identified in a total genome sequence so far (1). To explore the system of selenocysteine incorporation in B. subtilis, we screened serine-acceptable tRNAs to find an unknown tRNA for selenocysteine by the combined method of specific biotinylation of aa-tRNA (2) and RT-PCR (3). cDNAs obtained from the serine-acceptable tRNA pool were amplified and cloned into plasmid to read its sequence. This procedure gave cDNA library corresponding known serine tRNAs, but no candidate for selenocysteine has been found. Thus, this result, together with the previous data (4), might reveal that there is no selenocysteine tRNA in B. subtilis and/or metabolism of selenium is considerably different from known one as seen in other bacteria.
{"title":"A study of the method to pick up a selenocysteine tRNA in Bacillus subtilis.","authors":"J Matsugi, K Murao","doi":"10.1093/nass/44.1.149","DOIUrl":"https://doi.org/10.1093/nass/44.1.149","url":null,"abstract":"<p><p>In Bacillus subtilis, selenocysteine tRNA has not been identified in a total genome sequence so far (1). To explore the system of selenocysteine incorporation in B. subtilis, we screened serine-acceptable tRNAs to find an unknown tRNA for selenocysteine by the combined method of specific biotinylation of aa-tRNA (2) and RT-PCR (3). cDNAs obtained from the serine-acceptable tRNA pool were amplified and cloned into plasmid to read its sequence. This procedure gave cDNA library corresponding known serine tRNAs, but no candidate for selenocysteine has been found. Thus, this result, together with the previous data (4), might reveal that there is no selenocysteine tRNA in B. subtilis and/or metabolism of selenium is considerably different from known one as seen in other bacteria.</p>","PeriodicalId":19394,"journal":{"name":"Nucleic acids symposium series","volume":" 44","pages":"149-50"},"PeriodicalIF":0.0,"publicationDate":"2000-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1093/nass/44.1.149","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"22518009","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Y Yamamoto, Y Noutoshi, M Fujie, S Usami, T Yamada
To elucidate the contribution of LINE-like retrotransposon Zepps in formation and maintenance of chromosomal telomeres, newly formed mini-chromosomes in irradiated Chlorella vulgaris cells were isolated and structurally characterized. A mini-chromosome Y32 (approximately 400 kbp in size) was shown to have several copies of Zepp elements on both termini. On the right arm terminus, two copies of Zepps were found in a tandem array with poly(A) tracts facing towards the chromosome end. The poly(A) tail and a 3'-end of approximately 400 bp of the distal copy was replaced by telomeric repeats. On 5'-side of the proximal copy was another Zepp element in a reversed orientation. This newly formed telomeric structure is very similar to that found in the left arm terminus of chromosome I and support the model of Zepp-mediated maintenance of Chlorella telomeres.
{"title":"Analysis of double-strand-break repair by Chlorella retrotransposon Zepp.","authors":"Y Yamamoto, Y Noutoshi, M Fujie, S Usami, T Yamada","doi":"10.1093/nass/44.1.101","DOIUrl":"https://doi.org/10.1093/nass/44.1.101","url":null,"abstract":"<p><p>To elucidate the contribution of LINE-like retrotransposon Zepps in formation and maintenance of chromosomal telomeres, newly formed mini-chromosomes in irradiated Chlorella vulgaris cells were isolated and structurally characterized. A mini-chromosome Y32 (approximately 400 kbp in size) was shown to have several copies of Zepp elements on both termini. On the right arm terminus, two copies of Zepps were found in a tandem array with poly(A) tracts facing towards the chromosome end. The poly(A) tail and a 3'-end of approximately 400 bp of the distal copy was replaced by telomeric repeats. On 5'-side of the proximal copy was another Zepp element in a reversed orientation. This newly formed telomeric structure is very similar to that found in the left arm terminus of chromosome I and support the model of Zepp-mediated maintenance of Chlorella telomeres.</p>","PeriodicalId":19394,"journal":{"name":"Nucleic acids symposium series","volume":" 44","pages":"101-2"},"PeriodicalIF":0.0,"publicationDate":"2000-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1093/nass/44.1.101","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"22518138","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
M Ushioda, M Kadokura, Y Murami, T Moriguti, T Wada, K Seio, M Sekine
In this study, the solid-phase synthesis of oligodeoxyribonucleotides having a guanosine pyrophosphate cap structure (Gpp-) was achieved by using a new guanosine monophosphate unit having the DMTr group capable of estimation of coupling efficiency of the pyrophosphate bond formation. Since 7-methylguanosine base was unstable under basic conditions, Gpp-capped DNA oligomers were synthesized by using a new linker having a silanediyl bond, which allowed to release the DNA chain from the solid support by treatment with fluoride anion under neutral conditions.
{"title":"Solid-phase synthesis of deoxyribonucleotides having a cap structure analog.","authors":"M Ushioda, M Kadokura, Y Murami, T Moriguti, T Wada, K Seio, M Sekine","doi":"10.1093/nass/44.1.123","DOIUrl":"https://doi.org/10.1093/nass/44.1.123","url":null,"abstract":"<p><p>In this study, the solid-phase synthesis of oligodeoxyribonucleotides having a guanosine pyrophosphate cap structure (Gpp-) was achieved by using a new guanosine monophosphate unit having the DMTr group capable of estimation of coupling efficiency of the pyrophosphate bond formation. Since 7-methylguanosine base was unstable under basic conditions, Gpp-capped DNA oligomers were synthesized by using a new linker having a silanediyl bond, which allowed to release the DNA chain from the solid support by treatment with fluoride anion under neutral conditions.</p>","PeriodicalId":19394,"journal":{"name":"Nucleic acids symposium series","volume":" 44","pages":"123-4"},"PeriodicalIF":0.0,"publicationDate":"2000-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1093/nass/44.1.123","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"22518149","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
We have designed a new type of a DNA dendrimer which has rigid branched structure. The branching molecule was prepared from 1,3,5-tribromobenzene. The dendrimer unit, in which three oligonucleotide-chains, two molecules of T15 and one molecule of A15, linked to the branching molecule, was synthesized by an automated DNA synthesizer. The properties of the dendrimer unit and dendrimer formation by inter-molecular association of T15 and A15 chains of the dendrimer unit will be presented.
{"title":"Synthesis and properties of a new type DNA dendrimer.","authors":"Y Suzuki, T Otomo, H Ozaki, H Sawai","doi":"10.1093/nass/44.1.125","DOIUrl":"https://doi.org/10.1093/nass/44.1.125","url":null,"abstract":"<p><p>We have designed a new type of a DNA dendrimer which has rigid branched structure. The branching molecule was prepared from 1,3,5-tribromobenzene. The dendrimer unit, in which three oligonucleotide-chains, two molecules of T15 and one molecule of A15, linked to the branching molecule, was synthesized by an automated DNA synthesizer. The properties of the dendrimer unit and dendrimer formation by inter-molecular association of T15 and A15 chains of the dendrimer unit will be presented.</p>","PeriodicalId":19394,"journal":{"name":"Nucleic acids symposium series","volume":" 44","pages":"125-6"},"PeriodicalIF":0.0,"publicationDate":"2000-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1093/nass/44.1.125","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"22518150","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
K Shinozuka, N Matsumoto, A Nakamura, H Hayashi, H Sawai
A facile stereospecific synthetic method for alpha-anomeric 2'-deoxypyrimidine nucleoside unit utilizing aminooxazoline derivative of ribofuranose was investigated. Thus, easily accessible riboaminooxazoline derivative prepared by ribose and cyanamid was allowed to react with ethyl alpha-bromoethylacrylate to give corresponding adduct. The adduct was cyclized by strong base such as potassium t-butokiside. The resulted 2,2'-cyclonucleoside was then treated with acetyl bromide followed by n-butyltin hydride to give alpha-anomeric 3',5'-di-O-acetylthymidine. 3',5'-Di-O-acety groups of the nucleoside were easily removed by the action of excess of triethyl amine in methanol. Essentially same procedure afforded corresponding 2'-deoxyuridine, which was further, converted to alpha-anomeric 2'-deoxycytidine.
研究了一种利用核呋喃糖氨基恶唑啉衍生物立体定向合成α - 2′-脱氧嘧啶核苷元的简便方法。因此,由核糖和氰酰胺制备的易获得的核糖氨基恶唑啉衍生物可与α -溴乙基丙烯酸酯反应生成相应的加合物。加合物被t-丁托基酸钾等强碱环化。得到的2,2'-环核苷再用乙酰溴和正丁基氢化锡处理,得到α - 3',5'-二- o -乙酰胸苷。在甲醇中过量的三乙胺的作用下,核苷的3′,5′-二氧基很容易被去除。基本相同的过程得到相应的2'-脱氧尿苷,进一步转化为α - 2'-脱氧胞苷。
{"title":"Stereospecific synthesis of alpha-anomeric pyrimidine nucleoside.","authors":"K Shinozuka, N Matsumoto, A Nakamura, H Hayashi, H Sawai","doi":"10.1093/nass/44.1.21","DOIUrl":"https://doi.org/10.1093/nass/44.1.21","url":null,"abstract":"<p><p>A facile stereospecific synthetic method for alpha-anomeric 2'-deoxypyrimidine nucleoside unit utilizing aminooxazoline derivative of ribofuranose was investigated. Thus, easily accessible riboaminooxazoline derivative prepared by ribose and cyanamid was allowed to react with ethyl alpha-bromoethylacrylate to give corresponding adduct. The adduct was cyclized by strong base such as potassium t-butokiside. The resulted 2,2'-cyclonucleoside was then treated with acetyl bromide followed by n-butyltin hydride to give alpha-anomeric 3',5'-di-O-acetylthymidine. 3',5'-Di-O-acety groups of the nucleoside were easily removed by the action of excess of triethyl amine in methanol. Essentially same procedure afforded corresponding 2'-deoxyuridine, which was further, converted to alpha-anomeric 2'-deoxycytidine.</p>","PeriodicalId":19394,"journal":{"name":"Nucleic acids symposium series","volume":" 44","pages":"21-2"},"PeriodicalIF":0.0,"publicationDate":"2000-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1093/nass/44.1.21","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"22518381","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
H Maruoka, S Kitaoka, N Tohnai, Y Inaki, T Hatae, T Tanabe
Isopoly(S-carboxymethyl-L-cysteine) derivatives of nucleic acid bases were prepared as antisense compounds. These compounds in vitro have been found to form stable complex with oligo-DNA or RNA. This paper deals with effect of antisense compounds in vivo. The target in this paper is the sequence of the PSD-95 protein linked with NMDA receptor. Excess passing of calcium ions through the loss of the signal pathway without PSD-95 proteins caused by antisense compound. The cells detailing with L-cysteine derivatives showed the lowest percentage of 19.1%. The data were compared with that of phosphotioate antisense compound.
{"title":"The behavior and effect of isopoly (S-carboxymethyl-L-cysteine) derivatives of nucleic acid bases.","authors":"H Maruoka, S Kitaoka, N Tohnai, Y Inaki, T Hatae, T Tanabe","doi":"10.1093/nass/44.1.195","DOIUrl":"https://doi.org/10.1093/nass/44.1.195","url":null,"abstract":"<p><p>Isopoly(S-carboxymethyl-L-cysteine) derivatives of nucleic acid bases were prepared as antisense compounds. These compounds in vitro have been found to form stable complex with oligo-DNA or RNA. This paper deals with effect of antisense compounds in vivo. The target in this paper is the sequence of the PSD-95 protein linked with NMDA receptor. Excess passing of calcium ions through the loss of the signal pathway without PSD-95 proteins caused by antisense compound. The cells detailing with L-cysteine derivatives showed the lowest percentage of 19.1%. The data were compared with that of phosphotioate antisense compound.</p>","PeriodicalId":19394,"journal":{"name":"Nucleic acids symposium series","volume":" 44","pages":"195-6"},"PeriodicalIF":0.0,"publicationDate":"2000-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1093/nass/44.1.195","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"22518388","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
M Kitano, N Miyano-Kurosaki, Y Endo, M Yukita, H Takeuchi, Y Tamura, K Takai, M Nashimoto, H Takaku
We examined the suppression of virus expression by cleavage of the HIV-1 RNA gene using a mammalian tRNA 3' processing endoribonuclease and an External Guide Sequence Oligozyme (EGS) in vivo. We constructed an EGS expression vector that used the tRNA(met) promoter as an expression cassette for EGS. The EGS expression vector was targeted to the upstream region of gag, region. The EGS expression vector was co-transfected into COS cells with the HIV-1 gene plasmid vector. As compared with the EGS non-expressing cells and the EGS expressing cells, the EGS expressing cells with the targeted gag start codon had a clearly decreased amount of the HIV-1 gag p24 protein. The EGS expressing cells with the targeted gag start codon showed effective suppression of HIV-1 gene expression. Thus, these studies describe novel gene targeting agents for the inhibition of gene expression and antiviral activity.
{"title":"Effective suppression of HIV-1 gene expression by a mammalian tRNA 3' processing endoribonuclease and external guide sequence oligozymes.","authors":"M Kitano, N Miyano-Kurosaki, Y Endo, M Yukita, H Takeuchi, Y Tamura, K Takai, M Nashimoto, H Takaku","doi":"10.1093/nass/44.1.207","DOIUrl":"10.1093/nass/44.1.207","url":null,"abstract":"<p><p>We examined the suppression of virus expression by cleavage of the HIV-1 RNA gene using a mammalian tRNA 3' processing endoribonuclease and an External Guide Sequence Oligozyme (EGS) in vivo. We constructed an EGS expression vector that used the tRNA(met) promoter as an expression cassette for EGS. The EGS expression vector was targeted to the upstream region of gag, region. The EGS expression vector was co-transfected into COS cells with the HIV-1 gene plasmid vector. As compared with the EGS non-expressing cells and the EGS expressing cells, the EGS expressing cells with the targeted gag start codon had a clearly decreased amount of the HIV-1 gag p24 protein. The EGS expressing cells with the targeted gag start codon showed effective suppression of HIV-1 gene expression. Thus, these studies describe novel gene targeting agents for the inhibition of gene expression and antiviral activity.</p>","PeriodicalId":19394,"journal":{"name":"Nucleic acids symposium series","volume":" 44","pages":"207-8"},"PeriodicalIF":0.0,"publicationDate":"2000-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1093/nass/44.1.207","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"22518394","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Y Tanaka, T Hori, M Tagaya, M Katahira, F Nishikawa, T Sakamoto, Y Kurihara, S Nishikawa, S Uesugi
Three variants of minimized hepatitis delta virus (HDV) RNA ribozyme systems designed on the basis of the "pseudoknot" model were synthesized and their tertiary interactions were analyzed by NMR spectroscopy. Rz-1 is a cis-acting ribozyme system (the cleaved form, 56-mer) in which stem IV is deleted from the active domain of genomic HDV RNA. Rz-1 was uniformly labeled with stable isotopes, 13C and 15N. Rz-2 is a trans-acting ribozyme system (substrate: 8-mer, the cytidine residue at the cleavage site is replaced by 2'-O-methylcytidine; enzyme: 16-mer plus 35-mer). Rz-2 was partially labeled with stable isotopes in guanosine residues of enzyme 35mer. Rz-4 is a trans-acting ribozyme system (substrate: 8mer, the cytidine residue at the cleavage site is replaced by 2'-O-methylcytidine; enzyme 53mer) which was designed by Perrotta and Been. Rz-4 has the same sequence and an extra loop closing stem IV. From 2D-NOESY and 2D-HSQC (except for Rz-4) spectra, it was suggested each ribozyme forms "pseudoknot" type structure in solution. Additionally, it was found that G38 of Rz-1, G28 and G29 of Rz-2 and Rz-4 form base-pairs. These novel base-pairs are observed in the crystal structure of a modified genomic HDV RNA. From temperature change experiment of Rz-2, the imino proton signal of G28 disappeared at 50 degrees C earlier than the other corresponding signals. Upon MgCl2 titration of Rz-2, this signal showed the largest shift.
合成了基于“伪结”模型设计的三种最小化丁型肝炎病毒(HDV) RNA核酶系统变体,并利用核磁共振光谱分析了它们的三级相互作用。Rz-1是一种顺式核酶系统(裂解形式,56-mer),其中茎IV从基因组HDV RNA的活性区域中删除。Rz-1用稳定同位素13C和15N均匀标记。Rz-2是一种反式作用核酶体系(底物:8-mer,裂解位点的胞苷残基被2'- o -甲基胞苷取代;酶:16聚合体加35聚合体)。Rz-2在35mer酶的鸟苷残基中用稳定同位素进行了部分标记。Rz-4是一种反式作用核酶体系(底物:8mer,裂解位点的胞苷残基被2′- o -甲基胞苷取代;酶53mer),由Perrotta和Been设计。Rz-4具有相同的序列和一个额外的闭环干IV。从2D-NOESY和2D-HSQC(除了Rz-4)光谱中可以看出,每种核酶在溶液中形成“假结”型结构。此外,Rz-1的G38、Rz-2和Rz-4的G28和G29形成碱基对。这些新的碱基对在修饰的基因组HDV RNA的晶体结构中被观察到。从Rz-2的温度变化实验来看,G28的亚质子信号在50℃时比其他相应信号消失得更早。当Rz-2用MgCl2滴定时,信号变化最大。
{"title":"NMR analysis of tertiary interactions in HDV ribozymes.","authors":"Y Tanaka, T Hori, M Tagaya, M Katahira, F Nishikawa, T Sakamoto, Y Kurihara, S Nishikawa, S Uesugi","doi":"10.1093/nass/44.1.285","DOIUrl":"https://doi.org/10.1093/nass/44.1.285","url":null,"abstract":"<p><p>Three variants of minimized hepatitis delta virus (HDV) RNA ribozyme systems designed on the basis of the \"pseudoknot\" model were synthesized and their tertiary interactions were analyzed by NMR spectroscopy. Rz-1 is a cis-acting ribozyme system (the cleaved form, 56-mer) in which stem IV is deleted from the active domain of genomic HDV RNA. Rz-1 was uniformly labeled with stable isotopes, 13C and 15N. Rz-2 is a trans-acting ribozyme system (substrate: 8-mer, the cytidine residue at the cleavage site is replaced by 2'-O-methylcytidine; enzyme: 16-mer plus 35-mer). Rz-2 was partially labeled with stable isotopes in guanosine residues of enzyme 35mer. Rz-4 is a trans-acting ribozyme system (substrate: 8mer, the cytidine residue at the cleavage site is replaced by 2'-O-methylcytidine; enzyme 53mer) which was designed by Perrotta and Been. Rz-4 has the same sequence and an extra loop closing stem IV. From 2D-NOESY and 2D-HSQC (except for Rz-4) spectra, it was suggested each ribozyme forms \"pseudoknot\" type structure in solution. Additionally, it was found that G38 of Rz-1, G28 and G29 of Rz-2 and Rz-4 form base-pairs. These novel base-pairs are observed in the crystal structure of a modified genomic HDV RNA. From temperature change experiment of Rz-2, the imino proton signal of G28 disappeared at 50 degrees C earlier than the other corresponding signals. Upon MgCl2 titration of Rz-2, this signal showed the largest shift.</p>","PeriodicalId":19394,"journal":{"name":"Nucleic acids symposium series","volume":" 44","pages":"285-6"},"PeriodicalIF":0.0,"publicationDate":"2000-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1093/nass/44.1.285","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"22518604","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
DNA enzymes are RNA-cleaving single stranded DNA molecules. The structure and the catalytic domain of a DNA enzyme were determined by Santro et al. in 1997. In this study, we have designed several types of DNA enzymes (PB2Dz) targeted to the PB2 mRNA translation initiation region of influenza A virus, and examined their cleavage kinetics, nuclease resistance, and a luciferase gene reporter assay. Using a synthetic substrate, these DNA enzymes were shown to have cleavage activity that is dependent on the length of the substrate recognition domain. To confer serum nuclease resistance to the DNA enzymes, we designed a new type of DNA enzyme that has the N3'-P5' phosphoramidate modification (PB2Dz-N) at each terminal. We examined the activity of this DNA enzyme in vivo. The DNA enzymes used in this study inhibited the expression of the PB2-luciferase gene in COS cells. These results suggest that DNA enzymes are potentially useful as gene inactivating agents of influenza A virus.
{"title":"Inhibition of influenza virus RNA (PB2 mRNA) expression by a modified DNA enzyme.","authors":"H Takahashi, T Abe, K Takai, H Takaku","doi":"10.1093/nass/44.1.287","DOIUrl":"https://doi.org/10.1093/nass/44.1.287","url":null,"abstract":"<p><p>DNA enzymes are RNA-cleaving single stranded DNA molecules. The structure and the catalytic domain of a DNA enzyme were determined by Santro et al. in 1997. In this study, we have designed several types of DNA enzymes (PB2Dz) targeted to the PB2 mRNA translation initiation region of influenza A virus, and examined their cleavage kinetics, nuclease resistance, and a luciferase gene reporter assay. Using a synthetic substrate, these DNA enzymes were shown to have cleavage activity that is dependent on the length of the substrate recognition domain. To confer serum nuclease resistance to the DNA enzymes, we designed a new type of DNA enzyme that has the N3'-P5' phosphoramidate modification (PB2Dz-N) at each terminal. We examined the activity of this DNA enzyme in vivo. The DNA enzymes used in this study inhibited the expression of the PB2-luciferase gene in COS cells. These results suggest that DNA enzymes are potentially useful as gene inactivating agents of influenza A virus.</p>","PeriodicalId":19394,"journal":{"name":"Nucleic acids symposium series","volume":" 44","pages":"287-8"},"PeriodicalIF":0.0,"publicationDate":"2000-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1093/nass/44.1.287","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"22518605","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
A novel fluorophore viz. 4-dansylamido-1,8-naphthalimido-N-pentanol has been designed, prepared and characterised. The comparative fluorescence has been studied in different solvents, solvent gradient, aqueous solutions of inorganic ions and buffers. This can be used for covalent tagging of oligonucleotides having potential application in Molecular Biology.
{"title":"Novel fluorophore for labelling of oligonucleotides.","authors":"Y Singh, G Watal, K Misra","doi":"10.1093/nass/44.1.85","DOIUrl":"https://doi.org/10.1093/nass/44.1.85","url":null,"abstract":"<p><p>A novel fluorophore viz. 4-dansylamido-1,8-naphthalimido-N-pentanol has been designed, prepared and characterised. The comparative fluorescence has been studied in different solvents, solvent gradient, aqueous solutions of inorganic ions and buffers. This can be used for covalent tagging of oligonucleotides having potential application in Molecular Biology.</p>","PeriodicalId":19394,"journal":{"name":"Nucleic acids symposium series","volume":" 44","pages":"85-6"},"PeriodicalIF":0.0,"publicationDate":"2000-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1093/nass/44.1.85","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"22518705","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}