’ (Pharmacist-10) This was compounded by concerns over working with accuracy checking technicians (ACTs) ‘I’m a bit nervous…it’s still the pharmacist’s responsibility even though

it’s the ACT that has checked it.’ (Pharmacist-3). Essentially, pharmacists are taking on work unnecessarily whilst simultaneously disempowering their staff from taking responsibility for their work. This creates an impasse where neither pharmacist, staff or ultimately, customers benefit. Pharmacists delegate, but often incompletely; they also allow ‘reverse delegation’. Acknowledging that this behaviour potentially creates a workload problem Selleck BIBF1120 is essential. Better workload management could be achieved if pharmacists were only involved with tasks that specifically required them. Delegation could be a valuable tool in easing pharmacist workload pressures; effective FXR agonist staff planning and behaviour changes from the whole pharmacy team are requisites. Observation has given a unique insight into how effectively pharmacists delegate and manage their work albeit in a small sample of pharmacies. 1. Gidman W. Increasing community pharmacy workloads in England: causes and consequences. Int J Clin Pharm 2011; 33:

512–520. 2. Bond C, Blenkinsopp A, Inch J, Celino G, Gray, N. The effect of the new community pharmacy contract on the community pharmacy workforce. The Pharmacy Practice Research Unoprostone Trust 2008:1–34. Rachel Urban1,2, Nooresameen Rana1, Evgenia Paloumpi1, Julie Morgan1 1University of Bradford, Bradford, UK, 2Bradford Institute For Health Research,

Bradford, UK, 3Bradford Teaching Hospitals NHS Foundation Trust, Bradford, UK To determine which health care providers (HCPs) communicate with community pharmacy regarding changes to patients’ medication using semi-structured interviews. Community pharmacies receive information regarding changes to patients’ medication infrequently and inconsistently. Communication to community pharmacies in England must be increased to improve seamless care and reduce medication errors. Lack of communication to community pharmacy is a longstanding issue. Recently measures to improve communication have been introduced including guidance from the Royal Pharmaceutical Society (RPS)1 and the introduction the Discharge Medicines Review (DMR) service in Wales. Previous studies have shown that communication with community pharmacies can contribute toward effective, seamless care and reduce error, 2 however, there is little evidence which examines the range of different HCPs who currently liaise with community pharmacy. This study explored which HCPs communicate with community pharmacies regarding medication changes, the extent of the communication and solutions for improvement.

6b). Intriguingly, protected bands included the SMAG repeat labeled as c in Fig. 6b. The same result was obtained in RNA extension experiments, in which bands of elongation extended over SMAG repeat c only (Fig. 6c). We hypothesize that repeats a and

b fold into one large secondary structure, which is cleaved, and this promotes rapid 3′–5′ degradation of upstream 4478 transcripts. The number of predicted SLSs is significantly higher in prokaryotic genomes existing in nature than in random sequences of comparable GC content (Petrillo et al., 2006). This implies that the ability of a variety of sequences to fold into secondary structures is positively selected in prokaryotic genomes and may have functional significance. A fraction of SLSs is represented by REPs, http://www.selleckchem.com/screening/chemical-library.html sequences shown or hypothesized find protocol to serve different functions. REPs are binding sites for the integration host factor, a protein required for site-specific recombination and DNA replication

(Engelhorn et al., 1995). REPs are targets for the DNA gyrase (Espéli & Boccard, 1997), and repeats located between convergent genes may be a privileged target for the enzyme, in order to counteract the excess of positive supercoiling induced in the chromosome by DNA transcription (Moulin et al., 2005). As RNA elements, REPs may enhance the stability of 5′ proximal mRNA segments (Khemici & Carpousis, 2004). Finally, REPs induce innate immune system stimulation via TLR9, and could play a key role in the pathogenesis of Gram-negative septic shock (Magnusson et al., 2007). Tobes & Ramos (2005) established that, for a palindromic sequence to be considered as REP, the following criteria should be met: (a) be extragenic, (b) range in size from 21 to 65 bp and (c) constitute >0.5% of the total intergenic space. SMAGs meet all these criteria, and constitute the largest set of REPs described so far. SMAGs correspond to the repeats identified by Nunvar et

al. (2010). SMAGs can be sorted into five distinct subfamilies, CYTH4 and come in different genomic formats. Single units make up only 1/5 of the SMAG family. The remaining elements are organized as dimers or are grouped in tandem arrays of variable lengths. Altogether, SMAGs and intermingled DNA occupy 13% of the overall intergenic space, and make up 1.4% of the total chromosome. SMAG families residing in the environmental R551-3 and SKA14 S. maltophilia strains are comparable in size to the repeat family found in K279a. Yet, the sizes of some subfamilies vary, and K279a is enriched in SMAG-3. Most SMAG-3 are organized as HH dimers that feature conserved spacers, and may thus represent a relatively young sequence family variant. Changes in the abundance and chromosomal distribution may make SMAG-3 sequences suitable for use in accurate genotyping and epidemiological studies. Also, the ∼500 REPs identified in the E. coli MG1655 strain have been sorted into subfamilies.

Previous studies have demonstrated the influential Ruxolitinib datasheet role of striatal dopamine

levels on the locomotor response to a novel environment; for example, animals can be separated into two groups (high and low responders) according to their locomotor activity in reaction to a novel environment. Ferris et al. (2013) recently demonstrated that high and low response to novelty can predict both the tolerance that develops to cocaine directly at the dopamine transporter as well as the rate of acquisition of cocaine self-administration. Low novelty responders have been shown to have lower extracellular dopamine levels, in line with the present study, where we observed reduced functional activity in dopaminergic regions following 48 h withdrawal from cocaine self-administration (Verheij & Cools, 2008; Verheij et al., 2008). Previous work on withdrawal from cocaine self-administration has found depression of locomotor activity during similar time-points, an effect that is indicative of reductions in dopamine levels and ventral tegmental area cell firing (Gauvin et al., 1997; Koeltzow & White, 2003). Thus, the lower levels of locomotor activity would be predicted based on the reduced functional activity in dopaminergic nuclei. These alterations suggest that there could be changes in reward processing at baseline, which could play an important role in the reinstatement of cocaine-seeking (Schmidt

& Pierce, 2010). Because these regions are involved in the processing of salient stimuli, these data also suggest that the processing of alternative rewards, such as Dasatinib research buy food, may also be impaired at baseline (Carelli, 2002; Schultz, 2010). In addition, there may be differential effects of cocaine on dopaminergic systems involved in motor and reward processing, an effect that was highlighted in the behavioral data, indicating that there was no difference in baseline forward locomotion following cocaine self-administration. Note that there were reductions in stereotypic behaviors, indicating that although forward locomotion does not differ between groups, Cediranib (AZD2171) there may be inherent differences in motor control

following cocaine self-administration, although it is not clear as to the meaning of these results. Together, these data suggest that the alterations in functional activity are not general changes that occur in all dopaminergic terminal fields, but rather are specific to those associated with reward and reinforcement and selective aspects of motor control. In line with reductions in functional activity in dopaminergic regions 48 h following cocaine self-administration, electrophysiological recordings have demonstrated reduced action potential firing in nucleus accumbens neurons both in vitro and in vivo after withdrawal from cocaine self-administration (White et al., 1995; Dong et al., 2006; Ishikawa et al., 2009; Kourrich & Thomas, 2009; Mu et al., 2010).

difficile (Levett, 1986). This study was supported in part by the Slovenian Research Agency Grants J4-2236 and P4-0092). We thank Dr John Pringle, SLU, for critical reading of the manuscript. “
“Flexirubins are specific polyene pigments produced by several genera of Bacteroidetes. Colonies and cell extracts of Flavobacterium johnsoniae and Flexibacter elegans have been

investigated by Raman spectroscopy to show that this fast and non-destructive technique can be used to differentiate click here these pigments from carotenoids and to compare the flexirubin content of the two microorganisms. The presence or absence of certain distinguishing features in the CH combination band region at 2500–2750 cm−1 can assist in the discrimination between the two flexirubins investigated. Raman spectroscopy is thus a suitable selleck screening library tool not only to detect flexirubin pigments in bacterial cells, but also to further

characterize the pigments present in members of the Bacteroidetes genera that are rich in flexirubins. “
“Myxococcus xanthus has a large number of histidine kinase (HK) signal transduction proteins and many of these HKs are important for fruiting body development. Nla6S is an uncharacterized HK that lacks many of the conserved sequence motifs of typical HK proteins. In this study, we report that expression of the nla6S gene increases about sixfold during fruiting body development, that the Nla6S protein has the in vitro properties of HKs and that Nla6S is the prototype for a new family of HKs. To date, these Nla6-like HKs are found

only in fruiting members of the Cystobacterineae suborder of the myxobacteria. The myxobacterium Myxococcus xanthus has a highly social lifestyle. To obtain nutrients, gliding swarms of M. xanthus cells hunt prey bacteria and feed on them. When they are starving, M. xanthus cells initiate Megestrol Acetate a development cycle that yields multicellular fruiting bodies containing thousands of stress-resistant spores. Because of this multicellular lifestyle, M. xanthus has developed intricate signal transduction networks that monitor cell–cell signals and signals from the environment, and respond accordingly. Myxococcus xanthus has an abundance of histidine kinase (HK) sensor proteins to monitor these signals (Goldman et al., 2006). HKs, together with response regulators (RR), form a signal relay system known as the two-component signal transduction system (TCS). In this system, the HK autophosphorylates when it detects a particular signal and transfers the phosphoryl group to the RR, which activates it (Laub & Goulian, 2007). Activated RRs then alter the appropriate cellular process, often by modulating changes in gene expression. HKs typically contain a sensor and a transmitter domain (Stewart, 2010). The amino acid sequences of sensor domains are highly variable owing to the vast diversity of signals that they detect.

Individually, Gottron’s papules were seen in 91% (51/56) and heliotrope rash in 73% (36/49). Nailfold capillaroscopy

abnormalities were reported in 26 of 38 patients (68%). Calcinosis was not present in any patient at diagnosis (0/13); however, 18% (8/45) of patients with JDM had calcinosis documented during the course of the disease. Forty-four percent of chronic course patients (7/16) developed calcinosis compared with 4% of monophasic patients (1/21). No patient with polyphasic disease developed calcinosis. Dysphonia was documented in 14 patients and dysphagia in 11 patients at time of diagnosis. Throughout the course of the illness, 21 of 49 patients (43%) in whom there was adequate documentation had dysphonia and/or dysphagia. At presentation, arthritis buy BIBF 1120 was reported in 15 of 43 patients (35%) and Akt inhibitor contractures in 17 of 29 (59%). Of those

patients with contractures at onset, only five (29%) also had arthritis. Table 2 outlines the results of common investigations performed in the cohort. CK was the most frequently ordered muscle enzyme investigation (100% of patients) and was abnormal 65% of the time (37/57). Twenty patients had normal CK; four of these had no other enzyme measured and 16 had at least one other enzyme and this was abnormal in all cases. Aldolase was measured in only 10 patients and was abnormal in all. When measured, lactate dehydrogenase (LDH), aspartate aminotransferase (AST) and alanine aminotransferase (ALT) were abnormal 92% (23/25), 88% (29/33) and 58% (29/33) of the time, respectively. Two or more muscle enzymes were elevated in 65% of patients (37/57). Four patients (with only CK measured) had no abnormality

in muscle enzymes. All four demonstrated clinical weakness and supportive evidence of myositis with abnormal MRI, EMG or muscle biopsy. Erythrocyte sedimentation Acetophenone rate (ESR) was elevated in 84% (46/55) of patients. Muscle biopsy was performed in 29 patients and was abnormal in 83% (24/29). EMG was performed on four patients and was abnormal in all patients. Figure 2 outlines the frequency of use of muscle biopsy, EMG and MRI in the diagnostic work-up of patients over the period studied. MRI was performed on a total of 29 patients and demonstrated signs of myositis in 97% (28/29). One patient with normal MRI had treatment with oral steroids prior to the MRI. Antinuclear antibodies (ANA) were tested in 52 patients and titres were abnormal (titre > 1 : 160) in 33 (63%) cases. High titre antibody to extractable nuclear antigen (ENA) was detected in only one patient (1/32, 3%) and was directed toward topoisomerase-I. Table 3 outlines therapy at diagnosis and throughout the disease course of the cohort. Fifty-one percent (29/57) of patients were treated with steroids alone (oral and/or high-dose pulsed methylprednisolone) at diagnosis, of whom 12 (20%) received this as their only treatment throughout their disease course. High-dose pulsed intravenous steroids were used in a total of 47 (82%) patients.

, 2007). This notion led us to predict an important role for H 89 concentration any lipolytic enzyme of P. aeruginosa, which, like EstA, may have access to lipids of the bacterial outer membrane. Therefore, we have analysed the physiological role of the newly described lipase LipC, which also exerted significant effects on cellular motility as well as on the production of rhamnolipids. Accordingly,

biofilms formed by the lipC mutant showed a significantly different architecture than the corresponding wild-type biofilms. Rhamnolipids are detergent-like sugarlipids that may act as ‘wetting’ agents and also play a role as virulence factors (Daniels et al., 2004; Zulianello et al., 2006). The rhamnolipid biosynthesis pathway includes two sequential rhamnosyl transferase reactions (Rahim et al., 2001) starting from HHAs as precursors (Deziel et al., 2003), which are also present in culture supernatants and possess detergent-like properties (Deziel

et al., 2003). Recent studies have shown that HAAs as well as di-rhamnolipids can act as antagonizing stimuli on swarming motility (Tremblay et al., 2007). Rhamnolipids also play multiple roles in the maturation of biofilms because they promote motility and the maintenance of water-filled channels (Davey et al., 2003). Recently, experimental evidence was presented Tanespimycin solubility dmso indicating that twitching motility also requires rhamnolipid production. In the lipC mutant, swimming was also affected, whereas an rhlA mutant

did not show any difference as compared with the wild-type strain (data not shown). This result clearly indicates that the reduction in rhamnolipid DNA ligase production itself cannot explain the pleiotropic phenotype of the lipC mutant. Recently, Hancock’s lab has performed a comprehensive study on swarming motility of P. aeruginossa. They found that transposon insertion into a gene encoding the pseudopilus protein XcpU required for type II secretion resulted in decreased swarming motility and biofilm formation. However, it remained unclear whether XcpU itself exerted the observed effects or other secreted factors were also involved (Overhage et al., 2007). The swarming defect we have observed for the lipC mutant indeed indicates the requirement of additional extracellular enzymes as LipC has been shown to be secreted by the Xcp machinery (Martinez et al., 1999). Furthermore, two secreted lipolytic enzymes also interfere with motility in P. aeruginosa: (1) the autotransporter EstA located in the outer membrane is required for all types of motility and the formation of the typical architecture of wild-type biofilms and (2) the extracellular phospholipase PlcB is involved in twitching motility along phospholipid gradients (Barker et al., 2004), but its influence on swimming, swarming and biofilm formation is unknown.

Real-time PCR and chromatin immunoprecipitation analysis were then used to explore alterations in gene expression and modifications

Smad inhibitor of chromatin structure associated with the plastic outcome caused by fluoxetine in the visual system. Local infusion of 5-HT into visual cortex restored susceptibility to monocular deprivation in adulthood whereas infusion of WAY-100635, trkB-IgG or U0126 prevented the process of plasticity reactivation in fluoxetine-treated animals. Long-term fluoxetine treatment promoted a transient increase of Bdnf expression in the visual cortex, which was paralleled by an increased histone acetylation status at Bdnf promoter regions and by decreased expression of Hdac5. Accordingly, enhancing histone acetylation levels by systemic treatment with Trichostatin-A reactivated plasticity in the adult while

WAY-100635-infusion prevented epigenetic modifications in Bdnf promoter areas. The data suggest a key role for 5-HT1A receptor and BDNF-trkB signalling in driving a transitory epigenetic remodelling of chromatin structure that underlies the reactivation of plasticity in the visual system. “
“Gamma-band activity (30–90 Hz) and the synchronization of neural activity in the gamma-frequency range have been observed in different cortical and subcortical find protocol structures and have been associated with different cognitive functions. However, it is still unknown whether gamma-band synchronization subserves a single universal function or a diversity of functions across the full spectrum of cognitive processes. Here, we address this question reviewing the mechanisms of gamma-band oscillation generation and the functions associated with gamma-band activity across several cortical and subcortical structures. Additionally, we raise a plausible explanation of why gamma rhythms are found so ubiquitously across brain structures. Gamma band activity originates from the interplay between inhibition and excitation. We stress that gamma oscillations, associated with this interplay, originate

from basic functional motifs that conferred advantages oxyclozanide for low-level system processing and multiple cognitive functions throughout evolution. We illustrate the multifunctionality of gamma-band activity by considering its role in neural systems for perception, selective attention, memory, motivation and behavioral control. We conclude that gamma-band oscillations support multiple cognitive processes, rather than a single one, which, however, can be traced back to a limited set of circuit motifs which are found universally across species and brain structures. “
“To investigate the role(s) of protein-tyrosine sulfation in the retina, we examined retinal function and structure in mice lacking tyrosylprotein sulfotransferases (TPST) 1 and 2. Tpst double knockout (DKO; Tpst1−/−/Tpst2 −/−) retinas had drastically reduced electroretinographic responses, although their photoreceptors exhibited normal responses in single cell recordings.

Treatment limits progression of retinitis and reduces the risk of blinding complications such as retinal

detachment and macular involvement of CMVR [8]. Systemic anti-CMV treatment also provides prophylaxis to an unaffected contralateral eye. Intravitreal injections or implants containing anti-CMV treatment provide more expedient loading dosages if required and are localized treatments for those patients unable Bortezomib purchase to tolerate systemic therapy. Treatment of CMVR consists of an induction period of between 2 and 4 weeks of therapy followed by a maintenance period in which the drug dosage is lower. The duration of maintenance therapy depends on immune recovery with HAART and lack of evidence of CMVR progression learn more or reactivation. In a randomized study published by the Valganciclovir Study Group, the median time to progression of CMVR was 125 days for patients originally assigned to intravenous ganciclovir and 160 days for patients originally assigned to oral valganciclovir. The

proportions of patients in each group having a satisfactory response to induction therapy were similar between the two drugs, as were the rates of adverse events [7]. Systemic anti-CMV therapy should be considered as the first-line treatment strategy for CMVR (category 1 recommendation). The standard treatment regimens used in induction include oral valganciclovir 900 mg bd, iv ganciclovir 5 mg/kg bd, iv foscarnet 90 mg/kg bd. Intravenous cidofovir (5 mg/kg) is given weekly for 2 weeks. All intravenous dosages need adjustment in cases of renal impairment. Close monitoring for adverse events is required as anti-CMV medications may cause significant toxicities such as renal and electrolyte abnormalities, and bone marrow suppression. The additional use of a ganciclovir implant or intravitreal injections of ganciclovir/foscarnet is recommended for CMVR affecting zone 1 (see Fig. 5.1) [9]. Induction and maintenance with a ganciclovir implant should be considered in patients for whom systemic therapy is contraindicated. The median time to progression

of CMVR with a ganciclovir implant was approximately 220 days in the pre-HAART era [9]. The standard treatment regimens used in maintenance include oral valganciclovir 900 mg od, iv ganciclovir 5 mg/kg daily or 6 mg/kg/day for 5 days of the week, iv foscarnet 90 mg/kg od daily or 120 mg/kg for 5 GPX6 days of the week. Intravenous cidofovir (5 mg/kg) is given fortnightly. CMVR can be expected to relapse in spite of ongoing anti-CMV treatment if immune reconstitution does not occur [7]. Maintenance treatment can be stopped if there is good immune reconstitution (CD4>100 cells/μL and undetectable viral loads) [10–13]. This decision should be made following careful discussion between the HIV physician and the ophthalmologist involved in the patient’s care. When disease occurs in zones 1 and 2 (see Fig. 5.1), induction is achieved with oral valganciclovir as above.

These genomes vary in size from 18 844 nucleotides (nt) in Hanseniaspora uvarum to 109 103 nt in Moniliophthora perniciosa. There is no apparent correlation of genome size and gene content: size differences can be attributed to the size of introns and intergenic regions and the presence of integrated plasmids. Of the two major types of mitochondrial introns, type I is the norm in fungal mitochondrial genomes, while type II are usually present only in plant mitochondrial genomes (Lang et al., 2007). Trametes cingulata (Bakshi et al., 1970)

HIF inhibitor is a heterothallic dikaryon originally isolated from rotting Shorea robusta lumber. We present the sequence of the T. cingulata mitochondrial genome and compare it with the mitochondrial genomes of five basidiomycete species. Trametes is a representative genus of the polypore clade in the subphylum Agaricomycotina (Ko & Jung, 1999; Hibbett et al., 2007). The available mitochondrial

genomes of Basidiomycota at NCBI are represented by four species of Agaricomycotina including Pleurotus ostreatus (Wang et al., 2008), M. perniciosa (Formighieri et al., 2008), Schizophyllum commune and Cryptococcus neoformans var. grubii. The Ustilaginomycotina Atezolizumab clinical trial is a sister clade of the Agaricomycotina and we have selected Ustilago maydis as a representative for this group. Trametes cingulata was obtained from the American Type Culture Collection (http://www.atcc.org accession number ) and maintained and grown on 2% malt extract agar plates at room temperature. DNA was isolated from hyphae essentially as described by Raeder & Broda (1985). Hyphae were

collected by filtration or centrifugation, Abiraterone ic50 washed with 20 mM EDTA, pH 8.0, and freeze-dried for 24–48 h. Samples were crushed at room temperature in a mortar and resuspended in extraction buffer (200 mM Tris-Cl, pH 8.5, 250 mM NaCl, 25 mM EDTA, 0.5% SDS) using about 2 mL per 0.1 g of dried tissue. Phenol (∼0.7 vol.) was added to the slurry, which was then mixed for 2 min. Following the addition of ∼0.3 vol. chloroform, mixing and centrifugation at 10 000 g for 1 h, the aqueous layer was transferred to a new tube and 1/20 vol. of 20 mg mL−1 RNAse A was added and incubated 37 °C for 20 min. The RNAse was extracted with 1 vol. chloroform and the tube was centrifuged at 10 000 g for 10 min. DNA was precipitated from the aqueous layer by the slow addition of isopropanol (∼1 vol.). The precipitated mass of DNA was sequentially washed with 50% isopropanol and 70% ethanol, dried briefly and resuspended in TE buffer (10 mM Tris-Cl pH 7.5, 1 mM EDTA). This preparation included genomic and mtDNA. DNA was sequenced using a GLS FLX sequencer (http://www.454.com) and assembled using a gs de novo assembler (version 1.1.03). The final mitochondrial genome assembly was performed using bioinformatic procedures developed at the Computational Genetics Laboratory at the Minnesota Supercomputing Institute. The raw end reads of the assembled contigs were compared using blast (Altschul et al.

These genomes vary in size from 18 844 nucleotides (nt) in Hanseniaspora uvarum to 109 103 nt in Moniliophthora perniciosa. There is no apparent correlation of genome size and gene content: size differences can be attributed to the size of introns and intergenic regions and the presence of integrated plasmids. Of the two major types of mitochondrial introns, type I is the norm in fungal mitochondrial genomes, while type II are usually present only in plant mitochondrial genomes (Lang et al., 2007). Trametes cingulata (Bakshi et al., 1970)

CAL-101 nmr is a heterothallic dikaryon originally isolated from rotting Shorea robusta lumber. We present the sequence of the T. cingulata mitochondrial genome and compare it with the mitochondrial genomes of five basidiomycete species. Trametes is a representative genus of the polypore clade in the subphylum Agaricomycotina (Ko & Jung, 1999; Hibbett et al., 2007). The available mitochondrial

genomes of Basidiomycota at NCBI are represented by four species of Agaricomycotina including Pleurotus ostreatus (Wang et al., 2008), M. perniciosa (Formighieri et al., 2008), Schizophyllum commune and Cryptococcus neoformans var. grubii. The Ustilaginomycotina Selleckchem Navitoclax is a sister clade of the Agaricomycotina and we have selected Ustilago maydis as a representative for this group. Trametes cingulata was obtained from the American Type Culture Collection (http://www.atcc.org accession number ) and maintained and grown on 2% malt extract agar plates at room temperature. DNA was isolated from hyphae essentially as described by Raeder & Broda (1985). Hyphae were

collected by filtration or centrifugation, Tyrosine-protein kinase BLK washed with 20 mM EDTA, pH 8.0, and freeze-dried for 24–48 h. Samples were crushed at room temperature in a mortar and resuspended in extraction buffer (200 mM Tris-Cl, pH 8.5, 250 mM NaCl, 25 mM EDTA, 0.5% SDS) using about 2 mL per 0.1 g of dried tissue. Phenol (∼0.7 vol.) was added to the slurry, which was then mixed for 2 min. Following the addition of ∼0.3 vol. chloroform, mixing and centrifugation at 10 000 g for 1 h, the aqueous layer was transferred to a new tube and 1/20 vol. of 20 mg mL−1 RNAse A was added and incubated 37 °C for 20 min. The RNAse was extracted with 1 vol. chloroform and the tube was centrifuged at 10 000 g for 10 min. DNA was precipitated from the aqueous layer by the slow addition of isopropanol (∼1 vol.). The precipitated mass of DNA was sequentially washed with 50% isopropanol and 70% ethanol, dried briefly and resuspended in TE buffer (10 mM Tris-Cl pH 7.5, 1 mM EDTA). This preparation included genomic and mtDNA. DNA was sequenced using a GLS FLX sequencer (http://www.454.com) and assembled using a gs de novo assembler (version 1.1.03). The final mitochondrial genome assembly was performed using bioinformatic procedures developed at the Computational Genetics Laboratory at the Minnesota Supercomputing Institute. The raw end reads of the assembled contigs were compared using blast (Altschul et al.