A Tetrahymena thermophila G4-DNA Binding Protein with Dihydrolipoamide Dehydrogenase Activity

G4-DNA is a four-stranded structure that is formed by guanine-rich sequences. We report here the purification and characterization of a novel G4-DNA binding protein from Tetrahymena thermophila, designated TGP2. TGP2 was found to preferentially bind to G4-DNA oligonucleotides with adjacent single-stranded domains containing phosphorylated 5‘ ends and the sequence element, 5‘-ACTG-3‘. The amino acid sequence of TGP2 has high similarity to dihydrolipoamide dehydrogenase (DLDH) from a variety of species, and TGP2 was shown to have DLDH activity. Purified DLDH from porcine heart and bovine intestinal mucosa were shown to bind specifically to G4-DNA oligonucleotides. On the basis of these results we conclude that TGP2 is DLDH in T. thermophila and suggest that the G4-DNA binding capability of TGP2/DLDH may be biologically relevant

A Novel Uni-Acupoint Electroacupuncture Stimulation Method for Pain Relief

Electroacupuncture stimulation (EAS) has been demonstrated effective for pain relief and treating other various diseases. However, the conventional way of EAS, the bi-acupoint method, is not suitable for basis study of acupoint specificity. Moreover, its operations are inconvenient and difficult to be persevered, especially for long-term, continuous and even imperative treatments. These disadvantages motivate designs of new EAS methods. We present a novel uni-acupoint electrical stimulation method, which is applied at a single acupoint and quite meets the needs of basis study and simpler clinical application. Its pain relief effect has been evaluated by animal tests of Wistar rats. During the experiments, rats were given 30 min 2/100 Hz uni- and bi-acupoint EAS and their nociceptive thresholds before and after EAS were attained by hot-plate test. The analgesic effect was defined as the change of nociceptive threshold and used to evaluate the effectiveness of uni-acupoint EAS for pain relief. The hot-plate test results indicated that analgesic effect of uni-acupoint group was significantly higher than that of the control group and there was no significant difference of analgesic effects between uni- and bi-acupoint EAS. The results suggested that uni-acupoint method was an effective EAS method and had comparable pain relief effect with bi-acupoint method

Imaging Biological Samples with the Atomic-Force Microscope

The application of atomic force microscopy (AFM) to biological investigation is attractive for a number of reasons. Foremost among these is the ability of the AFM to image samples, even living cells, under near native conditions and at resolution equal to, or exceeding, that possible by the best light microscopes. Moreover, the ability of the AFM to manipulate samples it images provides a novel and far reaching application of this technology

A Tetrahymena thermophila G4-DNA Binding Protein with Dihydrolipoamide Dehydrogenase Activity

G4-DNA is a four-stranded structure that is formed by guanine-rich sequences. We report here the purification and characterization of a novel G4-DNA binding protein from Tetrahymena thermophila, designated TGP2. TGP2 was found to preferentially bind to G4-DNA oligonucleotides with adjacent single-stranded domains containing phosphorylated 5‘ ends and the sequence element, 5‘-ACTG-3‘. The amino acid sequence of TGP2 has high similarity to dihydrolipoamide dehydrogenase (DLDH) from a variety of species, and TGP2 was shown to have DLDH activity. Purified DLDH from porcine heart and bovine intestinal mucosa were shown to bind specifically to G4-DNA oligonucleotides. On the basis of these results we conclude that TGP2 is DLDH in T. thermophila and suggest that the G4-DNA binding capability of TGP2/DLDH may be biologically relevant.Reprinted (adapted) with permission from A Tetrahymena thermophila G4-DNA Binding Protein with Dihydrolipoamide Dehydrogenase Activity. Kehkooi Kee, Luming Niu, and Eric Henderson. Biochemistry 1998 37 (12), 4224-4234. DOI: 10.1021/bi9716377. Copyright 1998 American Chemical Society.</p

Atomic force microscopy of DNA-colloidal gold and DNA-protein complexes

The atomic force microscope (AFM)1 is capable of imaging and manipulating nucleic acids in solution and in air29' 13 We are developing methods for random and site-specific labeling of individual DNA molecules to facilitate manipulation of fragments excised in the AFM and for localization of specific DNA domains, such as protein binding sites and origins of replication. One successful method was to incorporate biotinylated nucleotides at random internal locations or specifically at the ends of linearized DNA molecules in vitro. Following complex formation with Snm diameter streptavidin-gold conjugates, chromatographic purification and passive adsorption of the complexes to mica, the biotinylated domains were easily localized in the AFM by virtue of the distinctive size and shape of the streptavidin-gold complex. In many cases unconjugated streptavidin (i.e., lacking gold) was also observed attached to the biotinylated DNA. A second approach to site-specific labeling of DNA for imaging in the AFM was to react DNA with restriction enzymes having sequence-specific binding properties. Like the unconjugated streptavidin-DNA complexes, these enzyme-DNA complexes were visible without attached colloidal gold. Efforts to image DNA labeled in vivo using bromodeoxyuridine (BrdU) and anti-BrdU antibodies are ongoing. Luming Niu ; Wenling Shaiu ; James Vesenka ; Drena D. Larson ; Eric Henderson; Atomic force microscopy of DNA-colloidal gold and DNA-protein complexes. Proc. SPIE 1891 (1993); doi:10.1117/12.146706.</p

Development of branchless watermelon near isogenic lines by marker assisted selection

Pruning is time-consuming and laborious in watermelon cultivation, which can not meet the needs for simplified cultivation in the future. The development of branchless lines will provide important germplasms for breeding watermelon varieties and is an important method for genetic improvement. In this study, the watermelon accession, Wu Cha Zao (WCZ) is a branchless inbred line that carries the branchless gene Clbl, which was used as the donor parent to develop branchless near isogenic lines (NILs). To construct the NILs of Clbl, WCZ crossed with the normal branching watermelon inbred line WT20 which was used as the recurrent parent. The co-segregating markers dCAPS10 and Indel1 with Clbl were used for foreground selection, and a total of 108 SSR markers was selected with good polymorphism between two parental lines for background selection which had relatively uniform distribution across 11 chromosomes. Using these markers to select individuals from the BC1F1, BC2F1, and BC2F2 generations, three NILs with a proportion of recurrent parent genome (PRPG) > 99% were finally obtained. The lateral branch and plant height phenotypes did not significantly differ between the NILs and WCZ, indicating that the NILs of Clbl under the genetic background of WT20 has been successfully developed. These results provide ideal materials for further in-depth analysis of the genetic mechanisms of lateral branch development and ideal plant architecture breeding in watermelon

Imaging Biological Samples with the Atomic-Force Microscope

The application of atomic force microscopy (AFM) to biological investigation is attractive for a number of reasons. Foremost among these is the ability of the AFM to image samples, even living cells, under near native conditions and at resolution equal to, or exceeding, that possible by the best light microscopes. Moreover, the ability of the AFM to manipulate samples it images provides a novel and far reaching application of this technology.This is a proceeding from 51st Annual Meeting of the Microscopy Society of America (1993): 512.</p