γPNA一种新型高效的肽核酸

肽核酸(peptide nucleic acid,PNA)是一种人工合成的具有类多肽骨架的DNA类似物,具有与核酸结合特异性强、组织和细胞内生物稳定性好、半衰期长等优点。通过靶向结合DNA/RNA而抑制其复制、转录和翻译过程,进行基因调控。在PNA骨架结构中γ位点引入带手性的官能团,能形成右手螺旋结构,显著提高其与靶DNA/RNA的杂交特性,这种PNA衍生物称之为γPNA。γPNA的溶解性、热稳定性和特异性等化学与生物学特性明显改善,在基因编辑和作为探针检测等方面具有良好的应用前景。通过对γPNA结构、性质及其研究进展进行总结,以期为γPNA反义应用提供理论依据和参考。     

肽核酸（PNA）的特性及其在分子生物学研究中的应用

ＰＮＡ是一类新的信息分子,具有不带电荷的类似于肽链的骨架结构并携带有碱基,能与互补的ＤＮＡ和ＲＮＡ分子杂交,故其在生物化学和分子生物学研究领域中具有广阔的应用前景。本文就其特性及其应用前景进行了简要叙述。     

表观遗传学（Epigenetics）

DNA的甲基化等修饰决定基因的表达。对这种不改变基因序列的表达调控的研究称为表观遗传学。它不仅是一项基础研究，还能揭示与癌症及生活习惯病的相关信息。东京大学的盐田邦郎教授为本期主讲。[编者按]     

卤虫中编码DM结构域的DNA序列的克隆和初步研究（简报）

性别决定的分子机制复杂多样,但是处于动物性别决定的基因调控网络底部的一些调控基因具有相当高的保守性。doublesex(dsx)基因和male abnomal-3(mab-3)基因分别是果蝇(Drosophila melanogaster)和线虫(Caenorhabditis elegans)性别决定调控途径末端的重要基因,对这两个基因序列的比较导致了DM结构域的发现,它是已知在性别发育过程中最为保守的DNA结合结构域。目前,已     

One new method of nucleic acid amplification—Loop-mediated isothermal amplification of DNA

Loop-mediated isothermal amplification (LAMP) is a novel nucleic acid amplification method, which amplifies DNA with high specificity, sensitivity, rapidity and efficiency under isothermal conditions using a set of four specially designed primers and a Bst DNA polymerase with strand displacement activity. The basic principle, characteristics, development of LAMP and its applications are summarized in this article.     

β乳球蛋白基因（βlg）的表达调控及其应用

β乳球蛋白（BLG）是反刍动物乳汁中的主要乳清蛋白,BLG表达受到βlg核心启动子,LCR,MAR等顺式作用元件和激素,转录因子等反式作用因子的调控,利用βlg启动子已在乳腺成功表达外源基因,但乳腺组织特异性表达外源基因时尚存在异位表达,差异表达,表达水平低和表达受位置效应影响等问题,构建表达载体时充分考虑βlg启动子和远端调控元件,有可能使上源基因获得,高效,特异的表达。     

小麦中外源基因瞬间表达调控研究及兔防御素（NP—1）基因的转化

将不同５上游调控序列驱动下的ＧＵＳ基因用基因枪法导入小麦幼胚和胚性愈伤组织,通过组织化学分析法和荧光分析法对ＧＵＳ基因的表达进行定量检测,比较了几种烟草花叶病毒（ＴＭＶ）Ω增强子序列对小麦中外源基因瞬间表达的调控作用;然后将其中效率最高的玉米Ｕｂｉｌ启动子与兔防御素（ＮＰ－１）连接起来,并加上Ｎｏｓ终止子,构民ＮＰ－１基因小麦表达载体,并转化小麦幼胚,经ＰＣＲ－Ｓｕｔｈｅｒｎ ｂｌｏｔ分析,初步确     

聚合酶链反应（Polymerase Chain Reaction，PCR）

PCR是80年代中期发展起来的体外核酸扩增技术。它具有特异、敏感、产率高、快速、简便、重复性好、易自动化等突出的优点;能在一个试管内将所要研究的目的基因或某一DNA片段于数小时内扩增至十万乃至百万倍,使肉眼能直接观察和判断;可从一根毛发、一滴血、甚至一个细胞中扩增出足量的DNA供分析研究和检测鉴定。过去几天几星期才能做到的事情,用PCR几小时便可完成。PCR技术是生物医学领域中的一项革命性创举和里程碑。     

果蝇前后图式基因调控的层次性（上）

一、引言由于分子遗传学的进步,对于基因如何代代相传以及个别基因的表达调控,我们已有相当的了解。然而对发育过程中,基因如何按一定的时空秩序,依次表达,并导致性状的发育,则所知甚少。这方面的研究还刚开始深入。已有人提出不同的理论模型,以解释在发     

Histone acetyltransferase p300 regulates the expression of human pituitary rumor transforming gene （hPTTG）

The human pituitary tumor transforming gene （hPTTG） serves as a marker for malignancy grading in several cancers, hPTTG is involved in multiple cellular pathways including cell transformation, apoptosis, DNA repair, genomic instability, mitotic control and angiogenesis induction. However, the molecular mechanisms underlying hPTTG regulation have not been fully explored. In this study, we found that overexpression of histone acetyltransferase （HAT） p300 upregulated hPTTG at the levels of promoter activity, mRNA and protein expression. Moreover, the HAT activity of p300 was critical for its regulatory function. Chromatin immunoprecipitation （CHIP） analysis revealed that overexpression of p300 elevated the level of histone H3 acetylation on the hPTTG promoter. Additionally, the NF-Y sites at the hPTTG promoter exhibited a synergistic effect on upregulation of hPTTG through interacting with p300. We also found that treatment of 293T cells with the histone deacetylase （HDAC） inhibitor Tfichostatin A （TSA） increased hPTTG promoter activity. Meanwhile, we provided evidence that HDAC3 decreased hPTTG promoter activity. These data implicate an important role of the histone acetylafion modification in the regulation of hPTTG.     

Peptide nucleic acids and the origin of life

The possibilities of pseudo-peptide-DNA mimics like PNA (peptide nucleic acid) having a role for the prebiotic origin of life prior to an RNA world is discussed on the basis of literature data showing that this type of molecules might have formed on the primitive earth (or other places in the universe), as well as data indicating the possibilities of template-directed PNA chemical replication and ligation. In particular, the merits of an achiral prebiotic genetic material is discussed.     

The first synthesis of chiral PNA monomer-cyclen conjugates.

A synthetic route to novel chiral PNA monomer-cyclen conjugates was described for the first time, the targeted products were obtained in high yields under mild reaction conditions. The preliminary results demonstrated that the uracil-PNA monomer-cyclen conjugates can rapidly bind Zn2+ in aqueous solution, and the structure of the Zn(II) complex was confirmed facilely by HRMS spectra, 1H NMR spectra and elemental analysis.     

Nucleobase‐caged peptide nucleic acids: PNA/PNA duplex destabilization and light‐triggered PNA/PNA recognition

The 2‐(o‐nitrophenyl)‐propyl (NPP) group is used as caging group to mask the nucleobases adenine and cytosine in N‐(2‐aminoethyl)glycine peptide nucleic acids (aeg‐PNA). The adeninyl and cytosinyl nucleo amino acid building blocks Fmoc‐a^NPP‐aeg‐OH and Fmoc‐c^NPP‐aeg‐OH were synthesized and incorporated into PNA sequences by Fmoc solid phase synthesis relying on high stability of the NPP nucleobase protecting group toward Fmoc‐cleavage, coupling, capping, and resin cleavage conditions. Removal of the nucleobase caging group was achieved by UV‐LED irradiation at 365 nm. The nucleobase caging groups provided sterical crowding effecting the Watson–Crick base pairing, and thereby, the PNA double strand stabilities. Duplex formation can completely be suppressed for complementary PNA containing caging groups in both strands. PNA/PNA recognition can be completely restored by UV light‐triggered release of the photolabile protecting group. Copyright © 2013 European Peptide Society and John Wiley & Sons, Ltd.     

The application of strand invasion phenomenon,directed by peptide nucleic acid (PNA) and single‐stranded DNA binding protein (SSB) for the recognition of specific sequences of human endogenous retroviral HERV‐W family

The HERV‐W family of human endogenous retroviruses represents a group of numerous sequences that show close similarity in genetic composition. It has been documented that some members of HERV‐W–derived expression products are supposed to play significant role in humans' pathology, such as multiple sclerosis or schizophrenia. Other members of the family are necessary to orchestrate physiological processes (eg, ERVWE1 coding syncytin‐1 that is engaged in syncytiotrophoblast formation). Therefore, an assay that would allow the recognition of particular form of HERV‐W members is highly desirable. A peptide nucleic acid (PNA)–mediated technique for the discrimination between multiple sclerosis‐associated retrovirus and ERVWE1 sequence has been developed. The assay uses a PNA probe that, being fully complementary to the ERVWE1 but not to multiple sclerosis‐associated retrovirus (MSRV) template, shows high selective potential. Single‐stranded DNA binding protein facilitates the PNA‐mediated, sequence‐specific formation of strand invasion complex and, consequently, local DNA unwinding. The target DNA may be then excluded from further analysis in any downstream process such as single‐stranded DNA‐specific exonuclease action. Finally, the reaction conditions have been optimized, and several PNA probes that are targeted toward distinct loci along whole HERV‐W env sequences have been evaluated. We believe that PNA/single‐stranded DNA binding protein–based application has the potential to selectively discriminate particular HERV‐W molecules as they are at least suspected to play pathogenic role in a broad range of medical conditions, from psycho‐neurologic disorders (multiple sclerosis and schizophrenia) and cancers (breast cancer) to that of an auto‐immunologic background (psoriasis and lupus erythematosus).     

Light-directed synthesis of peptide nucleic acids (PNAs) chips

We report herein the light-directed synthesis of peptide nucleic acids (PNAs) microarray using PNA monomers protected by photolabile protecting groups and a maskless technique that uses a digital micromirror array system to form virtual masks. An ultraviolet image from the virtual mask was cast onto the active surface of a glass substrate, which was mounted in a flow cell reaction chamber connected to a peptide synthesizer. Light exposure was followed by automatic chemical coupling cycles and these steps were repeated with different virtual masks to grow the desired PNA probes in a selected pattern. In a preliminary experiment, an array of PNA probes with dimensions of 4.11 mm × 4.11 mm was generated on each slide. Each synthesis region in the final array measured 210 μm × 210 μm for a total of 256 sites. The center-to-center space was 260 μm. It was observed from the hybridization pattern of the fluorescently labeled oligonucleotide targets that the fluorescence intensities of the matched, and mismatched sequences showed substantial difference, demonstrating specificity in the identification of complementary sequences. This opens the way to exploit processes from the microelectronics industry for the fabrication of PNA microarrays with high densities.     

Solid-phase synthesis of peptide nucleic acid (PNA) monomers and their oligomerization using disulphide anchoring linkers

A new simple solid-phase method has been developed for synthesizing Boc-protected peptide nucleic acid (PNA) monomers. An immobilized backbone 3 was built on Expansin® resin using an ester disulphide handle: 2-hydroxypropyl-dithio-2′-isobutyric acid (HPDI). The base acetic acids of thymine 5 , Z-cytosine 9 , Z-adenine 12 , and 6-O-benzyl guanine 17 were prepared and coupled to the immoblized backbone. The HPDI handle was cleaved under mild conditions by cyanolysis or assisted hydrolysis with tris(2-carboxyethyl)phosphine (TCEP) to give undamaged PNA monomers. These monomers were coupled to form oligomers by solid-phase method with another disulphide linkage: aminoethyldithio-2-isobutyric acid (AEDI) grafted on an amino-functionalized TentaGel® resin, using in situ neutralization and TBTU as activating reagent. Final cleavage of the AEDI linker gave PNA bearing a cysteamide residue that could be useful for optimizing PNA properties. Oligomers of up to 16 residues long were assembled. © 1998 European Peptide Society and John Wiley & Sons, Ltd.     

多肽核酸

通过计算机模型设计出一种以肽骨架取代ＤＮＡ中脱氧核糖磷酸酯骨架的ＤＮＡ类似物．实验已证明其与互补寡核苷酸的结合具有高度的特异性．综述了其合成、杂交性及在分子生物学等方面的应用．