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The data from the measurement on algae were globally fit to three

The data from the measurement on algae were globally fit to three exponential decays. This result suggested

Selleckchem 3-MA that the three lifetimes could be treated as separate pools of PSII that cannot transfer between each other. Two of the populations had lifetimes of 65 and 305 ps, with the third having a lifetime of 1 ns. The amplitudes of the two shorter lifetimes increased during the light treatment and decreased in the ensuing darkness. In addition, these amplitudes substantially decreased when the pH gradient was dissipated using nigericin. The amplitudes associated with the 65 and 305 ps lifetime components exhibited different dynamics during qE induction and relaxation, which led us to suggest that there are two different mechanisms associated with qE in C. reinhardtii. This technique correlates the T axis, which describes the timescales of qE triggering, with the t axis, which probes changes in the membrane and photophysical mechanism of qE. Fig. 10 Schematic of “fluorescence lifetime snapshots” measurements. The technique tracks changes on both the T timescale (sec to hours) as well as in the t timescale (ps to ns). qE triggering

and the thylakoid membrane rearrangement Avapritinib cost occur on the T timescale. Quenching of chlorophyll fluorescence occurs on the t timescale and contains information about the membrane configuration As discussed in the “Fluorescence lifetimes” section and Appendix B, the insight from fitting fluorescence lifetimes to multiple exponential decays is limited. Using the fluorescence

lifetime snapshot measurements to differentiate between different hypotheses for qE mechanisms requires fitting the fluorescence lifetimes to a detailed mechanistic model of energy transfer. Because different energy transfer models are able to fit fluorescence Ketotifen lifetime data well (van der Weij-de Wit et al. 2011), much theoretical and experimental progress remains to be made in developing accurate models of energy transfer in PSII. We are optimistic that future developments in this area will enable the interpretation of fluorescence lifetime snapshots in the context of a mechanistic model for qE. Concluding remarks Looking forward, much progress in the development of experimental techniques and theoretical models will be needed before the site(s) and mechanism(s) of qE are identified and the triggering processes and ensuing membrane changes are characterized. Obtaining unambiguous answers is particularly challenging because the pigments and proteins involved in qE are found inside of a lipid membrane, are PI3K Inhibitor high throughput screening buried within a cell, are highly dependent on interactions with their local environment, and undergo changes on a wide range of timescales.

enterocolitica WA or Y pestis Ind195 at MOI 1 and 20, respective

enterocolitica WA or Y. pestis Ind195 at MOI 1 and 20, respectively, for 1 h. Following stimulation with 10 ng/ml TNF-α at 5 h post-infection, luciferase activity was measured 24 h post-infection. Results were determined from two independent experiments performed in triplicate. A ‘*” denotes that the % NF-κβ inhibition using the inhibitors was significantly different (p<0.05) compared to the no drug control (black).

The relative NF-κB inhibition by Yersinia infection was determined as a percentage of luciferase Staurosporine ic50 activity in bacteria-infected cells relative to luciferase activity in bacteria-free control cells. (B) THP-1 cells were pretreated with the small JAK inhibitor molecules and infected with Y. enterocolitica WA or Y. pestis Ind195 at MOI 5 and 20, respectively, for 1 h. TNF-α levels were determined by ELISA on conditioned

media collected 24 h post-infection. Results were determined from two representative independent experiments Trichostatin A clinical trial performed in quadruplicate. A ‘*” denotes that TNF-α release using inhibitors was significantly different (p<0.05) compared to the no drug control. Cytokine release in response to purified LPS from E. coli 055:B5 (5μg/ml, light blue) was used as a control for pro-inflammatory mediator signaling. (C) Normal HDC were pre-treated with the small molecules for 18 h prior to infection with Y. enterocolitica WA or Y. pestis KIM5-. Bacterial infection was stopped 1 h post-infection with 170 μg/ml chloramphenicol. TNF-α levels were determined by ELISA on conditioned media collected 24 h post-infection. Statistical analysis was performed on data from 3 experiments performed in quadruplicate. TNF-α release in response to all inhibitor treatments were statistically significant (p<0.05) compared to no drug controls. We also tested the effect of the small molecule TBB, an inhibitor of the CKII Mirabegron serine

kinase, which functions in cell stress response, cell cycle and cell growth regulation by activation of IKK. CKII also regulates expression of HSPH1, another stress response gene identified in our shRNA screen [26]. Similar to OSI930, pretreatment of RE-luc2P-HEK293, THP-1, and NHDC cells with TBB resulted in higher levels of NF-κB-regulated gene expression and TNF-α release compared to a no drug control, in response to both Y. enterocolitica and Y. pestis infection (Figure 3A-C, blue vs black bars). The small molecule CKI-7 was used to validate the role of SGK1 (serum and glucocorticoid-inducible kinase 1) on NF-κB-regulated gene expression in response to Yersinia infection. SGK1 is a serine/threonine kinase that functions in cellular stress response and regulates activity of the epithelial sodium channel ENaC [27, 28], a function shared with WNK1, another kinase identified from the shRNA screen. Incubation of RE-luc2P-HEK293 cells with CKI-7 resulted in increased NF-κB-mediated luciferase activity upon exposure of Y. enterocolitica and Y. pestis-infected cells to TNF-α (Figure 3A, purple vs black bars).

Hence, all risk estimates are above 1 Largely, the ranking

Hence, all risk estimates are above 1. Largely, the ranking

according to adjusted PR estimates is in accordance with the ranking based on crude prevalence, with a few exceptions indicative of some confounding. After identifying three occupational subgroups with a relatively high risk of contact sensitisation to the thiurams, namely healthcare workers (physicians, nurses and related), food processors (cooks, meat and fish processors) and professional cleaners, the issue of a possible differential time trend was addressed. In view of (i) a distinct general risk gradient related to age (Table 2) and (ii) a weak, but significant association between age and year of patch test in the IVDK population (Uter et al. 2008), simple bivariate

Smoothened inhibitor analyses of crude sensitisation prevalence across time were avoided. Instead, three separate Poisson regression models including age as confounder and the year of patch test as exposure of interest were used to identify a significant decline of sensitisation prevalence in case of healthcare workers (p for trend = 0.0008), but no significant trend for the other two subgroups. The time course of age-standardised sensitisation prevalences is shown in Fig. 1a for healthcare workers and in Fig. 1b for the two other occupational groups. Fig. 1 a Time trend of sensitisation to the Selleckchem BIX 1294 thiuram mix in healthcare workers. Sensitisation prevalence is directly age standardised. Straight grey line LDN-193189 clinical trial represents the fitted regression line to represent a linear subgroup-specific trend. b Time trend of sensitisation to the thiuram mix in food handlers and cleaners, respectively. Sensitisation prevalence is directly age standardised. Straight grey lines represent fitted regression lines to represent a linear subgroup-specific trend Discussion Thiurams and dithiocarbamates, which are also represented by the thiuram mix in patch testing (Andersen

et al. 2006), are important constituents of natural and synthetic rubber products. The vulcanisers (accelerators) may occur both in occupational and non-occupational context (e.g., in privately used “household gloves” (Proksch et al. 2009)). A considerable amount of unreacted accelerator—be it thiurams or other classes—remains Oxaprozin in the cured rubber product, migrates to the surface and comes into contact with the skin. At least in thin products such as gloves or condoms, it is possible to reduce the residual amount, and, with it, dermal exposure, by washing with hot water to create a product, which is more or less “hypoallergenic” in this respect (Andersen et al. 2006). Although rubber products, in particular, rubber gloves, constitute the major part of dermal exposure, additional rather limited skin contact with thiurams may also be due (i) to pesticides (Saunders and Watkins 2001), (ii) fungicides, also in paints and (iii) to animal repellents (Andersen et al. 2006).

For example, if we wish to discern whether the biofilm is respond

For example, if we wish to discern whether the biofilm is responding to iron limitation, we first identify a set of genes that are up-regulated in response to iron deprivation (e.g. the work of Ochsner [9]). The rank of each of these transcripts in the biofilm data set is then compared to transcript ranks for the same genes in data sets collected from both rapidly

growing and deliberately iron-starved cultures. In this way it becomes possible to evaluate physiological activities in the biofilm rather than just documenting differences between the biofilm and a reference state. In the experiments reported here, RNA was extracted from an entire, homogenized biofilm specimen. An obvious concern with this approach is that it neglects the inherent biological Ro 61-8048 mw heterogeneity of the biofilm [1]. We would like to address this concern upfront with two points. First, just because a population is heterogeneous

does not mean that measurements of population averages are invalid. Population averages are very widely and informatively used in biology. Second, we suggest that even the concept of an average may not be appropriate in this case. The current conceptual model of P. aeruginosa drip-flow biofilms is that they consist of two distinct populations: an aerobic, metabolically active upper layer and a lower, and larger, layer consisting of inactive CX-5461 mouse cells containing very low levels of mRNA [10, 11]. Because the inactive cells contain so little RNA, this majority is expected to be essentially invisible on the microarray. From this perspective, the transcriptomes reported here may best be thought of as reflecting the properties of the transcriptionally-active subpopulation rather than the average behavior of the entire population. These concepts are AZ 628 mouse elaborated on in the Results Carnitine palmitoyltransferase II and Discussion. Results and Discussion Three day old drip flow biofilms of P. aeruginosa were characterized with respect to antibiotic tolerance, oxygen availability, and microscale patterns of protein synthetic activity. These biofilms

contained 9.56 ± 0.31 cfu cm-2. Reduced antibiotic susceptibility of biofilm bacteria P. aeruginosa cells grown in biofilms were protected from killing by tobramycin and ciprofloxacin, in comparison to actively growing planktonic bacteria. Both antibiotics rapidly and effectively reduced viable cell numbers in an aerobic, planktonic culture. After 12 h of treatment with 10 μg ml-1 tobramycin or 1.0 μg ml-1 ciprofloxacin, planktonic log reductions measured were 3.18 ± 1.79 (n = 3, ± SD) and 4.84 ± 0.55 (n = 3, ± SD) for tobramycin and ciprofloxacin, respectively. In contrast, neither antibiotic was very effective against biofilms of P. aeruginosa. After 12 h exposure to antibiotic in continuously flowing medium, the log reductions in viable cell numbers were 0.72 ± 0.56 (n = 3, ± SD) and 1.37 ± 0.06 (n = 3, ± SD) for tobramycin and ciprofloxacin, respectively.

2 v 4 9 months; P = 0 48), CNS progression or local brain tumor r

2 v 4.9 months; P = 0.48), CNS progression or local brain tumor response. (9.5 v 8.3 months; P = 0.95). None of those trials detected any benefit for theses end point mentioned above. In the trial by Mehta et al. [23], no difference in survival or time to neurological learn more progression was seen in the use of motexafin

gadolinium and WBRT versus WBRT alone. However, a subgroup analysis, carried out for lung cancer patients was reported to as an improvement in neurological progression favoring the motexafin gadolinium and WBRT arm. The results for the lung cancer subgroup can only be interpreted as a hypothesis generated as there was no a priori decision to analyze this group independently. On the basis of these results, a phase III trial was conducted exclusively in patients with NSCLC; a preliminary report was presented at the 2006 ASCO meeting. In this international trial, 554 patients were randomly assigned to WBRT (30 Gy in 10 fractions) plus MGd (5 mg/kg with each RT treatment) or WBRT alone [24]. There was a trend to an increased time to neurological progression, the primary endpoint in the study, in patients receiving

MGd (15.4 versus 10 months with RT alone). In another large RCT study [27], Suh et al. showed in a subset analysis that the addition of efaproxiral to WBRT reduced the death rate by 46% (P < 0.0086). Quality of life was improved in the WBRT with efaproxiral arm compared to the WBRT alone arm (P = 0.019). Quality-adjusted CB-839 nmr survival was statistically and significantly improved by the addition of efaproxiral to WBRT (P = 0.001). Patients with brain metastasis may suffer a certain degree Clomifene of JIB04 neurocognitive function (NCF) impairment from multiple factors including the tumor, WBRT, neurosurgical procedures, chemotherapy and other neurotoxic therapies (including anticonvulsants and steroids), or from paraneoplastic effects

induced by the malignancy [41]. Three trials included in this meta-analysis evaluated neurocognitive function. However, we were not able to pool these data, due to the different methods used for this outcome. In addition to that, studies involving NCF deterioration should be carefully interpreted. NCF decline in the literature is often defined statistically and there is little consensus as to the actual clinical relevance of a statistical definition. Conventionally, the measures used, such as the Folstein mini-mental status examination, are rather crude, and it is crucial to develop sensitive and practical neurocognitive function testing to characterize these changes [30]. In particular, the sensitivity of mini-mental status examination has been shown to be problematical in detecting subtle neurocognitive dysfunction in patients with brain metastasis where clinically apparent WBRT-induced dementia is rare (1.9–5.1%) [42, 43].

Clin

Clin selleck products Genet 2008, 73: 545–553.eFT508 concentration CrossRefPubMed 15. Tao H, Shinmura K, Suzuki M, Kono S, Mibu R, Tanaka M, Kakeji Y, Maehara Y, Okamura T, Ikejiri K, Futami K, Yasunami Y, Maekawa T, Takenaka K, Ichimiya H, Imaizumi N, Sugimura H: Association between genetic polymorphisms of the base excision repair gene MUTYH and increased colorectal cancer risk in a Japanese population. Cancer Sci 2008, 99: 355–360.CrossRefPubMed 16. Kasahara M, Osawa K, Yoshida K, Miyaishi A, Osawa Y, Inoue N, Tsutou A, Tabuchi Y, Tanaka K, Yamamoto M, Shimada E, Takahashi J: Association of MUTYH Gln324His and APEX1 Asp148Glu

with colorectal cancer and smoking in a Japanese population. J Exp Clin Cancer Res 2008, 27: 49.CrossRefPubMed 17. Barbone F, Bovenzi M, Cavallieri F, Stanta G: Cigarette smoking and histologic type of lung cancer in men. Chest 1997, 112 (6) : 1474–1479.CrossRefPubMed 18. Paz-Elizur T, Sevilya Z, Leitner-Dagan Y, Elinger D, Roisman LC, Livneh Z: DNA repair of oxidative DNA damage in human carcinogenesis: potential application for cancer risk assessment and prevention. Cancer Lett 2008, 266: 60–72.CrossRefPubMed 19. Al-Tassan N, Eisen T, Maynard J, Bridle H, Shah B, Fleischmann C, Sampson JR, Cheadle

JP, Houlston RS: Inherited variants in MYH are unlikely to contribute to the risk of lung carcinoma. Hum Genet 2004, 114: 207–210.CrossRefPubMed 20. Ali M, Kim H, Cleary S, Cupples C, Gallinger S, Bristow R: Characterization of mutant MUTYH proteins associated with CH5424802 in vitro familial colorectal cancer. Gastroenterology 2008, 135: 499–507.CrossRefPubMed 21. Toyokuni S, Mori T, Dizdaroglu M: DNA base modifications in renal chromatin of wistar rats treated with a renal carcinogen, ferric nitrilotriacetate. Int J Cancer 1994, 57: 123–128.CrossRefPubMed Competing interests The authors declare that they have no competing interests. Authors’ contributions AM, KO and JT plan the study made all coordination and was involved in the laboratory processing. YO, NI, KY and MK participated in the study and performed the statistical analysis. AT, YT, KS and NT carried out handling the samples. All authors read and approved the final version

of manuscript.”
“Background Colorectal Cytidine deaminase cancer (CRC) is one of the most common causes of cancer death throughout the world. Multistage development of the disease has been associated with remarkable genetic events, mainly at the level of oncogenes and oncosuppressor genes, most notably the adenomatous polyposis coli gene (APC) [1], ras [2, 3], and p53 [4]. Although great advances have been made during the last few decades in understanding the molecular biology of colorectal cancer [5], the prognosis of patients with this neoplasm has not improved in parallel. The overall five-year survival rate remains poor (40–45%) [6]. It can be assumed that several genes involved in the pathogenesis of colorectal cancer are still unknown.

430PubMedCrossRef 8 Meerburg BG, Singleton GR, Kijlstra A: Roden

430PubMedCrossRef 8. Meerburg BG, NCT-501 in vitro Singleton GR, Kijlstra A: Rodent-borne diseases and their risks for public health. Crit Rev Microbiol 2009,35(3):221–270.PubMedCrossRef 9. Vinetz JM: Leptospirosis. Curr Opin Infect Dis 2001,14(5):527–538.PubMedCrossRef 10. Mayer-Scholl A, Draeger A, Luge E, Ulrich R, Nockler K: Comparison of two PCR systems for the rapid detection of Leptospira spp. from kidney tissue. Curr Microbiol 2011,62(4):1104–1106.PubMedCrossRef learn more 11. Yang KJY, Luo YP, Wu GQ, Yang ZP, Kang

ZG: Epidemiology of leptospirosis in Liping county, Guizhou, 2001–2008. Dis Surveill 2009,24(10):768–769. 12. Morey RE, Galloway RL, Bragg SL, Steigerwalt AG, Mayer LW, Levett PN: Species-specific identification of Leptospiraceae by 16S rRNA gene sequencing. J Clin Microbiol 2006,44(10):3510–3516.PubMedCrossRef 13. Ahmed A, Thaipadungpanit J, Boonsilp S, Wuthiekanun V, Nalam K, Spratt BG, Aanensen DM, Smythe LD, Ahmed N, Feil EJ: Comparison of two multilocus sequence based genotyping

schemes for Leptospira species. PLoS Negl Trop Dis 2011,5(11):e1374.PubMedCrossRef 14. Romero EC, Blanco RM, Galloway RL: Analysis of multilocus sequence typing for identification of Leptospira isolates in Brazil. J Clin Microbiol CBL0137 ic50 2011,49(11):3940–3942.PubMedCrossRef 15. Caimi K, Varni V, Melendez Y, Koval A, Brihuega B, Ruybal P: A combined approach of VNTR and MLST analysis: improving molecular typing of Argentinean isolates of Leptospira interrogans. Memorias do Instituto Oswaldo Cruz 2012,107(5):644–651.PubMedCrossRef 16. Enright MC, Spratt BG: Multilocus sequence typing. Trends Microbiol 1999,7(12):482–487.PubMedCrossRef 17. Yalin W, Lingbing Z, Hongliang Y, Jianmin X, Xiangyan Z, Xiaokui G, Utpal P, Jinhong Q: High prevalence of pathogenic Leptospira in wild Florfenicol and domesticated

animals in an endemic area of China. Asian Pac J Trop Med 2011,4(11):841–845.PubMedCrossRef 18. Perez J, Brescia F, Becam J, Mauron C, Goarant C: Rodent abundance dynamics and leptospirosis carriage in an area of hyper-endemicity in New Caledonia. PLoS Negl Trop Dis 2011,5(10):e1361.PubMedCrossRef 19. Subharat S, Wilson PR, Heuer C, Collins-Emerson JM: Investigation of localisation of Leptospira spp. in uterine and fetal tissues of non-pregnant and pregnant farmed deer. N Z Vet J 2010,58(6):281–285.PubMedCrossRef 20. Faine SAB, Bloin C, Perolat P: Leptospira and leptospirosis. 2nd edition. Melbourne, Australia: MedSci; 1999. 21. Zhang CCNY, Li XW, Cui ZG, Jiang XG: Application of multiple-locus variable-number tandem repeat analysis (MLVA) for molecular typing of Leptospira interrogans serogroup lcterohaemorrhagiae. Chin J Microbiol Immunol 2009,29(12):1144–1147. 22. Guo SHDZ, Li JH: Analysis of leptospirosis epidemic in 31 provinces (1991–2005). J Public Health Prevent Med 2006, 6:8–10. 23. Yang M, Mo RJ: Exploration of Space Distribution on Leptospirosis Epidemic Focus with Host Animal. Practical Prevent Med 2007, 14:46–54.

The organic template moiety in the sample was determined using a

The organic template moiety in the sample was determined using a Mettler TGA SDTA851 instrument (Mettler-Toledo, Columbus, OH, USA) with a heating rate of 10°C·min−1 under nitrogen flow. Nitrogen adsorption-desorption analysis was conducted using a Micromeritics ASAP 2010 instrument (Norcross, GA, USA). The template-free PI3K Inhibitor Library sample was first degassed at 250°C for 3 h followed by

nitrogen adsorption measurement at −196°C. The surface physicochemical properties were then calculated using the Brunauer-Emmett-Teller (BET) and the Barrett-Joyner-Halenda (BJH) models [21]. Solid-state 29Si-MAS-NMR spectra were recorded using a Bruker Ultrashield 300 spectrometer (Madison, WI, USA) operating at 300 MHz with tetramethylsilane as a reference. The measurement

was carried out at 79.4 MHz and single-contact cross-polarization 4EGI-1 pulse program was used. The spectra were acquired with a pulse length of 2.7 μs, a repetition time of 6 s, and a contact time of 4 ms. The FTIR spectra of the as-synthesized solid products were obtained with a PerkinElmer spectrometer (System 2000) using the KBr pellet technique (KBr/sample weight ratio = 150:1). Results and discussion The chemical composition of the initial and re-used solutions characterized by dry mass, AAS, and TG/DTA analyses is summarized in Table  1. As can be seen, large amounts of silicate solution (approximately 15 g) and CTABr (approximately 3.5 g) were consumed for three subsequent synthesis cycles of MCM-41. Initially, the CTABr was dissolved in distilled water, and silica was precipitated out after sodium silicate was added into the CTABr solution. At this stage, silicate oligomers act as multidentate ligands with high charge density at head groups, which leads to a lamellar organization of the surfactant [22]. As the acid is introduced, polycondensation and polymerization of silica take place, resulting in the dissolution of lamellar phase. At pH close to 11.0, this dissolution is followed by the formation Gemcitabine datasheet of the hexagonal Ilomastat research buy MCM-41 material [22, 23]. Table 1 Compensated chemicals added into non-reacted mother liquor for MCM-41 synthesis

cycles and MCM-41 solid yield MCM-41 synthesis 1st cycle 2nd cycle 3rd cycle Non-reacted mother liquor (g) 0 54.404a 63.337a Added reagents Na2SiO3 (g) 21.206 15.664 15.560 CTABr (g) 5.772 3.750 3.251 H2O (g) 79.916 31.882 27.110 H2SO4 (g) 0.603 2.082 0.9881 pH 10.78 10.80 10.80 Solid yield, gram (wt.%)b 8.034 g (73.6%) 7.851 (71.9%) 7.694 (78.3%) aAfter evaporating water at 55°C for 16 h. b . pH was determined to be the most important of the investigated synthesis parameters in affecting pore ordering and mesophase. The solubility and the rate of dissolution of silica increases with the increasing pH resulted in a decrease of the total interfacial area and a more long-range pore ordering [24, 25]. High pH results in fast and complete hydrolysis where polymerization can occur within a few minutes [25].

Strain construction To construct strain NF33, a 400 bp DNA fragme

Strain construction To construct strain NF33, a 400 bp DNA fragment from the region upstream of B. cereus lysK was amplified by PCR using primers NF36F and NF36R, cut with EcoRI and BamHI, and ligated into

similarly restricted pDG268 [28] to produce the plasmid pBCJ307. pBCJ307 was inserted into the amyE locus of the B. subtilis lysine auxotroph strain 1A765 by double crossover to produce strain NF33. In order to analyze the effect of a reduction of the cellular level of charged tRNALys on expression of a P lysK(T box) lacZ fusion, strain BCJ367 was constructed. Plasmid pBCJ307 was integrated into the B. subtilis chromosome AZD1480 clinical trial by a double crossover event at the amyE locus to produce strain BCJ363. To place the endogenous lysS gene of B. subtilis under IPTG inducible control, plasmid pMUTIN4 [29] was digested with SalI and BsiWI and eluted from an agarose

gel Omipalisib purchase to remove the 2 kb lacZ gene. The ends of the plasmid molecule were blunt ended using Klenow polymerase and religated, resulting in plasmid pMUTINXZ. A 670 bp DNA fragment encoding the end of the yacF gene was amplified with oligonucleotides NF2F and NF2R using B. subtiliis strain 168 chromosomal DNA as a template. This fragment was digested with EcoRI and inserted into the EcoRI site of pMUTINXZ, resulting in plasmid pXZ2. Plasmid pXZ2 was then integrated onto the chromosome of strain BCJ363 by a Campbell enough type event check details generating strain BCJ366 thereby placing expression of the lysS gene under the control of the IPTG inducible Pspac promoter. To effect tight control of the Pspac promoter, replicating plasmid pMap65 [30] that encodes a lacI gene, was transformed into BCJ366 to produce strain BCJ367. Strain NF54 was made to assess whether a B. subtilis strain expressing a T-box regulated lysK gene was viable. A 1.95 kb fragment of the B. cereus chromosome encoding the lysK promoter, leader region and structural gene was

generated by PCR using oligonucleotides NF36F and NF9R. This fragment was digested with EcoRI and cloned into the EcoRI site of plasmid pBCJ102 that has transcriptional terminators flanking the multiple cloning site, to generate plasmid pNF30 [31]. A 2567 bp fragment encoding the lysK promoter, T box element and structural gene flanked by transcriptional terminator sequences was amplified using the pBluescript T7 and M13 reverse primers and plasmid pBCJ102 as template. The ends of this fragment were phosphoryalted using T4 polynucleotide kinase (Promega) and it was then cloned into the EcoRV site of plasmid pDG1730 [32] to produce the plasmid pNF48. Plasmid pNF48 was integrated at the amyE locus of the B. subtilis chromosome by a double crossover event to produce strain NF52.