Jinjiang, China
Jinjiang, China

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Zhang L.,Northwest University, China | Zhang L.,Longdong College | Jia J.,Northwest University, China | Xu Y.,Vanderbilt University | And 3 more authors.
In Vitro Cellular and Developmental Biology - Plant | Year: 2012

We have obtained transgenic plants of Nicotiana sylvestris expressing an arylalkylamine N-acetyltransferase (AANAT) gene and a hydroxyindole-O-methyltransferase (HIOMT) gene using Agrobacterium tumefaciens-mediated transformation. Both AANAT and HIOMT are key enzymes in melatonin synthesis. Transgenic plants of N. sylvestris were characterized by polymerase chain reaction (PCR) and reverse transcription PCR analyses. The content of melatonin was significantly higher in transgenic plants than in nontransgenic plants. The highest melatonin content of transgenic plant leaves reached 50. 4 μg/g dry weight, while almost no melatonin could be detected in the nontransformed plants. To investigate effects of the expression of the AANAT and HIOMT genes on melatonin function in plants, isolated protoplasts of N. sylvestris were exposed to ultraviolet (UV)-B radiation for different durations. DNA damage was evaluated by single-cell gel electrophoresis, which showed that the TailDNA percentage value in transgenic protoplasts was lower than that in the nontransformed protoplasts in the range of 0-30 s of UV-B radiation. DNA damage caused by UV-B was therefore reduced in transgenic N. sylvestris plants. © 2011 The Society for In Vitro Biology.

Zhang L.-J.,Northwest University, China | Zhang L.-J.,Longdong College | Hao J.-G.,Northwest University, China | Jia J.-F.,Northwest University, China
Plant Physiology Communications | Year: 2010

Protoplasts of Nicotiana sylvestris was used for experimental material to investigate DNA damage in protoplast which was induced by 0.5 W-cm-2 UV-B for different time (0, 5, 10, 30, 60, and 120 s) by using comet assay. The results showed that the indexes of tail moment and olive tail moment etc which detected the DNA damage had statistically dose effect relationships with the time of UV-B radiation from 0 to 10 s. Comet assay detected the degree of DNA damage of tobacco protoplast induced by UV-B radiation rapidly and sensitively.

Zhang L.-J.,Northwest University, China | Zhang L.-J.,Longdong College | Jia J.-F.,Northwest University, China | Hao J.-G.,Northwest University, China | And 2 more authors.
Mutation Research - Genetic Toxicology and Environmental Mutagenesis | Year: 2011

In the present study, we developed a modified protocol for the basic comet assay that increased efficiency without sacrificing assay reliability. A spreader was used to spread agarose-embedded cells on a slide, making the manipulation and processing of multiple samples easier. Using this technique, we are able to rapidly prepare five or more comet assay samples on one slide. To demonstrate the effect of the protocol modifications on assay reliability, we present an example of how the comet assay was used in our laboratory to analyze the effect of melatonin (N-acetyl-5-methoxitryptamine; MEL) on the DNA repair ability of Gentiana macrophylla Pall. protoplasts after irradiation with different doses of ultraviolet-B radiation. A slight, but statistically significant (P< 0.01), dose-related protective effect of MEL was observed in our experiments. The first use of the comet assay was to confirm the antioxidant and DNA repair functions of MEL in plants. The modified protocol is cost-effective and provides substantial advantages over the conventional comet assay. © 2011 Elsevier B.V.

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