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High efficiency method to obtain supercoiled DNA with a commercial plasmid purification kit

100%
Carbone A., Fioretti F. M., Fucci L., Ausió J., Piscopo M.
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2012
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vol. 59
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issue 2
275-278
EN
Supercoiled state corresponds to the active form for plasmid applications. The relaxed circular form of plasmids is often inactive or poorly active. To obtain significant amounts of almost fully supercoiled DNA, we modified the standard protocol of a commercially available Qiagen plasmid purification kit. Our changes led to isolation of almost 100% of the plasmids in the supercoiled state. The modified protocol was used to purify different plasmids with consistent results. The purified plasmids maintain supercoiled state for about two months. The modified protocol is very advantageous because it allows easy DNA production with high degree of supercoiled form at low cost.
2
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Self-association of Chaetopterus variopedatus sperm histone H1-like. Relevance of arginine content and possible physiological role

86%
Salvati D., Conforti S., Conte M., Matassa D. S., Fucci L., Piscopo M.
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2008
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vol. 55
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issue 4
701-706
EN
Self-association of histones H1 from calf thymus and from sperm of the marine worm Chaetopterus variopedatus was studied on native and glutaraldehyde cross-linked molecules by PAGE and by salt-induced turbidity measurements. Multiple polymers were generated by native sperm histone H1-like after glutaraldehyde cross-linking while the same treatment on its lysine- or arginine-modified derivatives and on somatic histone H1 failed to induce polymerization. This result suggests the relevance of arginine content in the formation of histone H1-like polymers particularly because Chaetopterus variopedatus and calf thymus histones H1 have similar content of lysine but different K/R ratio (2 and 15, respectively). Salt-induced turbidity experiments confirmed the high tendency of sperm histone H1-like to form oligomers, particularly in the presence of phosphate ions. Native PAGE analysis in the presence of phosphate supported this hypothesis. The reported results suggest that phosphate ions connecting lysine and arginine side chain groups contribute to the interaction of sperm histone H1-like with DNA in chromatin and play a key role in organization and stabilization of the chromatin higher order structures.
3
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H3K4 histone methylation in oral squamous cell carcinoma

86%
Mancuso M., Matassa D. S., Conte M., Colella G., Rana G., Fucci L., Piscopo M.
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issue 3
405-410
EN
Methylation of specific lysine residues in histone tails has been proposed to function as a stable epigenetic marker that directs biological functions altering chromatin structure. Recent findings have implicated alteration in heterochromatin formation as a contributing factor in cancer development. In order to verify whether changes in the overall level of H3K4 histone methylation could be involved in oral squamous carcinoma, the levels of H3K4me1, me2 and me3 were measured in oral squamous carcinoma, leukoplakias and normal tissues. The levels of H3K4me2 and me3 were significantly different in oral squamous cell carcinoma in comparison with normal tissue: the level of H3K4me2 was increased while that of H3K4me3 decreased. No significant differences could be found between the two types of tissues in the level of H3K4me1. A similar trend was found in the leukoplakias that appeared more like the pathological than normal tissue. These results support the idea that alteration of chromatin structure could contribute to oncogenic potential.
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