网站标志
商品搜索
您好!欢迎光临18587057888 
点评详情
发布于:2019-2-12 04:07:52  访问:2 次 回复:0 篇
版主管理 | 推荐 | 删除 | 删除并扣分
Foil and incubated 4? days at 42 . To observe the effects of photoreactivation
PubMed ID:https://www.ncbi.nlm.nih.gov/pubmed/26080418 IWR-1Biological Activity Sequences for Haloarcula marismortui, Natronomonas pharaonis, and Methanosphaera stadtmanae have been downloaded from NCBI (see More file 1). One blot was utilized to measure total harm (CPDs and 6-4PPs); the other was exposed to CPD photolyase and visible light to eliminate CPDs and let for detection of 6-4PPs alone. The blots were then exposed to rabbit polyclonal antiserum containing antibodies to 6-4PPs and CPDs, then to biotinylated anti-rabbit antibody followed by alkaline phosphatase-conjugated Extravidin (Sigma) and lastly to Nitro Blue tetrazolium (NBT) and 5-bromo-4-chloroindolyl phosphate (BCIP) substrate. The intensity of blue color was proportional towards the volume of DNA harm in the samples and was measured working with a scanning densitometer (BioRad GS-670) and compared to a set of requirements included on each blot.Foil and incubated four? days at 42 . To observe the effects of photoreactivation on survival, two identically spotted plates have been exposed to Sylvania GroLux fluorescent light (Sylvania F40/GRO/AQ/RP) for 24 hours before incubation at 42 . One plate was wrapped in foil as a control.Repair experiments and Immunoassays for measurement of photoproducts The repair experiments and dot-blot immunoassays for UV photoproducts had been carried out as described previously [12,15,21,41]. Log-phase cells were harvested and irradiated with 150 J/m2 UV-C at a dose rate of 1 J/m2/sec as well as the irradiated cells had been incubated aerobically at 37 to enable repair to proceed. We‘ve got previously shown that, just after this dose of UV, there‘s PubMed ID:https://www.ncbi.nlm.nih.gov/pubmed/25681438 no detectable DNA replication in the course of a 3-hour post-UV incubation [15]. All irradiation and post-UV incubation was carried out either below yellow light illumination or in the dark. Fifty-ml samples have been harvested at timed intervals and genomic DNA extracted utilizing Promega Wizard genomic DNA kits. DNA samples in the numerous time points were equalised by measuring fluorescence of ethidium bromide-stained DNA in agarose gels, adjusting DNA concentrations and repeating this evaluation as quite a few occasions as essential until all samples were of equal concentration.and 0322, respectively. PubMed ID:https://www.ncbi.nlm.nih.gov/pubmed/26080418 Sequences for Haloarcula marismortui, Natronomonas pharaonis, and Methanosphaera stadtmanae were downloaded from NCBI (see Extra file 1). Amino acid sequences have been aligned working with CLUSTAL_X1.83 [42]. Alignments have been manually inspected and edited if needed. TREEPUZZLE5.2 was applied for quartet puzzling consensus maximum likelihood phylogenetic reconstruction using the JTT amino acid substitution matrix [43].Competing interestsThe author(s) declare that they have no competing interests.Authors‘ contributionsDJC and SM conceived and designed the study and cowrote the manuscript. DJC and MC constructed the mutants and carried out preliminary characterization. ER, LS and IB confirmed mutant genotypes by PCR and Southern analysis; ER and LS performed UV survival experiments. IB carried out the repair experiments; SM carried out the immunoassays. BB and SD performed the phylogenetic analysis and assisted with preparation of the manuscript. All authors study and approved the final manuscript.Extra material Additional FileAccession numbers for phylogenetic analysis. Accession numbers utilized to generate data presented in Figure five.
共0篇回复 每页10篇 页次:1/1
共0篇回复 每页10篇 页次:1/1
我要回复
回复内容
验 证 码
看不清?更换一张
匿名发表 
帖子分类
脚注信息

---------------Copyright ©2011-2027 德国事务--------------All rights reserved-------------

------------------------请您扫描微信二维码添加专业德国事务顾问为好友------------------------

微信二维码

----------------------------------------------------------------------------------------------------