RESUMO
Here we demonstrate the use of multiple Si nanochannel (NC) or nanograting (NG) instead of the conventional single nanochannel or nanowire design in biosensors. The NG devices can significantly reduce device-to-device variation, and improve device performance, e.g. higher current, higher ON/OFF ratio, smaller subthreshold slope, lower threshold voltage Vt in buffer solution. NG devices also result in higher sensor stability in buffer and diluted human serum. We believe such improvements are due to reduced discrete dopant fluctuation in the Si nanowires and biochemical noise in the solution because of the multiple-channel design. The improved devices allow us to sense pH linearly with 3-aminopropyltriethoxysilane coated devices, and to selectively detect insulin with limit of detection down to 10 fM in both buffer solution and diluted human serum without pre-purification.
Assuntos
Técnicas Biossensoriais , Insulina/isolamento & purificação , Nanoestruturas/química , Humanos , Concentração de Íons de Hidrogênio , Insulina/sangue , Limite de Detecção , Nanofios/química , Propilaminas , Silanos/química , Silício/químicaRESUMO
p21(Waf1/Cip1) protein levels respond to DNA damage; p21 is induced after ionizing radiation, but degraded after UV. p21 degradation after UV is necessary for optimal DNA repair, presumably because p21 inhibits nucleotide excision repair by blocking proliferating cell nuclear antigen (PCNA). Because p21 also inhibits DNA mismatch repair (MMR), we investigated how p21 levels respond to DNA alkylation by N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), which triggers the MMR system. We show that MNNG caused rapid degradation of p21, and this involved the ubiquitin ligase Cdt2 and the proteasome. p21 degradation further required MSH2 but not MLH1. p21 mutants that cannot bind PCNA or cannot be ubiquitinated were resistant to MNNG. MNNG induced the formation of PCNA complexes with MSH6 and Cdt2. Finally, when p21 degradation was blocked, MNNG treatment resulted in reduced recruitment of MMR proteins to chromatin. This study describes a novel pathway that removes p21 to allow cells to efficiently activate the MMR system.
Assuntos
Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Reparo do DNA/efeitos dos fármacos , Metilnitronitrosoguanidina/farmacologia , Proteína 2 Homóloga a MutS/metabolismo , Proteínas Nucleares/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitinação/efeitos dos fármacos , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Cromatina/genética , Cromatina/metabolismo , Inibidor de Quinase Dependente de Ciclina p21/genética , Dano ao DNA/efeitos dos fármacos , Dano ao DNA/fisiologia , Reparo do DNA/fisiologia , Células HeLa , Humanos , Proteína 1 Homóloga a MutL , Proteína 2 Homóloga a MutS/genética , Proteínas Nucleares/genética , Antígeno Nuclear de Célula em Proliferação/genética , Antígeno Nuclear de Célula em Proliferação/metabolismo , Complexo de Endopeptidases do Proteassoma/genética , Complexo de Endopeptidases do Proteassoma/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitinação/fisiologiaRESUMO
Little is known about the significance of insects as environmental reservoirs of antibiotic-resistant bacteria. We characterized the antibiotic resistome of the microbial community in gypsy moth larval midguts by applying functional metagenomics to cultured isolates. The minimum inhibitory concentrations of 12 antibiotics were determined for 44 cultured isolates, and antibiotic resistance genes were selected from metagenomic libraries derived from DNA extracted from a pool of the isolates. Six unique clones were identified. Two were highly resistant to penicillin-type beta-lactams, two were moderately resistant to erythromycin, and two were moderately resistant to a range of antibiotics, including erythromycin, carbenicillin, and chloramphenicol. Sequence analysis predicted that the active genes encoded efflux pumps, a transcriptional activator of efflux pump protein expression, and an extended-spectrum class A beta-lactamase. Insect guts are a reservoir of antibiotic resistance genes with the potential for dissemination.
