4. Aftereffects of Disasters: Grief, Traumatic Grief, and ResiliencyFaculty, staff, and other diverse populations have long reported the scope and http://www.selleckchem.com/products/Vandetanib.html magnitude of grief up to and including traumatic grief following a range of disasters (e.g., natural, human-made, etc.). For example, O’Mallon [18] stated, ��Global events such as earthquakes, tsunamis, the 9/11 attack, and Hurricane Katrina fall into the category of traumatic grief. Grief associated with loss of life, home, and finances left families vulnerable ���� (paragraph 12). Although the authors of the current study agree that disasters can engender traumatic grief, we do not believe that human beings are fated to experience traumatic grief following disasters. Nonetheless, the empirical research suggests that the possibility exits.

Thus, it is important to understand the variability and types of grief reported and how the grief process may relate to coping after disasters. Toward this end, K��bler-Ross’ [19] five stages of grief may provide a framework to make meaning of the aftereffects and outcomes often reported by individuals who experience hurricanes, disasters, and other traumatic and adverse events. K��bler-Ross’ [19] introduced five stages of grief (i.e., denial, anger, bargaining, depression, and acceptance) following death. Overtime, the five stages of grief have also been used to explain the range of reactions and possibly the ��phases�� that people move through following various losses (e.g., natural disasters, death, loss of income, and divorce, to name a few). Stage 1 (Denial).

Denial leads to initial comments such as ��No, not me, it cannot be true�� [19, page 33]. Denial is usually a ��temporary defense�� and transition into ��partial acceptance�� [19, page 34]. During denial, the initial shock caused by the loss is experienced. Stage 2 (Anger). According to K��bler-Ross, denial ��is replaced by feelings of anger, rage, envy, and resentment�� (page 43). At this stage, the grieving person questions ��Why me?�� (page 43). He or she is ���� very difficult to cope with from the point of view of family and staff �� anger is displaced in all directions and projected onto the environment at times almost at random�� (page 43). Stage 3 (Bargaining).

K��bler-Ross illustrated the grieving person’s mentality during this stage as ��If we have been unable to face the sad facts in the first period and have been angry at people and God in the second phase, maybe we can succeed in entering Cilengitide into some sort of an agreement which may postpone the inevitable happening�� (page 71). K��bler-Ross also described a sense of guilt among the grieving during this stage. Stage 4 (Depression). According to K��bler-Ross [19], when the illness or issue cannot be ignored, ���� numbness or stoicism �� anger and rage will soon be replaced with a sense of great loss�� (page 75).

..4. Discussion4.1. Roosting Behavior in Mustached BatsThis study is the first to show that in mustached bats, individuals roost at highly restricted locations within a colony. Whether this is also true when individuals roost at locations outside the colony is less clear. The roosting locations of males overlap very little, at least in the short term. These roosting locations most likely drive http://www.selleckchem.com/products/crenolanib-cp-868596.html several social interactions that take place in the dark where visual cues are absent, but olfactory and auditory cues are abundant. Conspecifics most likely maintain their territories by scent marks made by rubbing the anogenital region against the substrate. Accordingly, crouching is likely required to monitor the scent boundaries.

In our observations, marking behavior sometimes alternated with crouching, which was consistent with the idea that these behaviors are related. Our observation that reintroduced bats increase their marking and crouching behavior is also evidence that these behaviors helped to establish and confirm roost position. Scent marking of territories with various exudates has been reported in several bat species [33, 34], including marking with the anogenital region for at least four species [20, 35].4.2. Acoustic Signal Design in Mustached BatsThis study demonstrated a strong association between calls and discrete behavior patterns in the mustached bat (see Table 1). A tight temporal binding between calls and behaviors may be especially important for communication in species, such as mustached bats, that roost in a completely dark environment.

Two noisy calls, rBNB and fSFM, and their composites were associated with two agonistic behaviors, nipping, and fighting. This association is consistent with the Motivation-Structure hypothesis, which posits that animals motivated by aggression will produce relatively low frequency, noisy, broadband calls [24, 31]. The level of noisiness in the fSFM call may correspond to the intensity of aggression, but this remains to be tested. In previous observations of mustached bats in a cage, playback of rBNB startled or warned an approaching bat and even made it turn back [26]. Other calls failed to elicit a similar response. In addition to the bro
Although the prevention science approach that focuses on risk and protective factors of high-risk adolescent behavior has generated much research and prevention programs in the past few decades, it has been criticized Drug_discovery as focusing too much on adolescent problems and pathology. As such, there is an alternative approach that emphasizes the importance of positive youth development.

2.1. Measurement MethodologyMeasurements of wind farm construction noise were performed from a drifting rigid hull inflatable boat (RHIB) in the vicinity of the piling site [7]. To avoid interaction with the hydrophone, the engine, Belinostat HDAC radar, and echosounder were turned off. The geographic position and time of measurement were recorded with a handheld GPS GARMIN GPSMap60 at a frequency of one position every 5 seconds. The clock of the recorder was synchronised beforehand with the GPS-time (UTC). At the start and the end of each measurement a reference signal was recorded. Several recordings of few minutes each (1 to 5min.) were performed at different locations on September 26 2009 (monopile A02) and January 15 2010 (monopile B10) at the Blighbank and on the May 11 (jacket CG3) and the of July 12 2011 (jacket CB6) at the Thorntonbank site (Table 1).

Weather conditions encountered during fieldwork featured a wind force of 1�C3BF and a sea state of 1 to 2.Table 1Geographic position, peak level (Lz�Cp), and distance from the piling location of the underwater noise measurements at the Blighbank site (monopiles A02 and B10) and at the Thorntonbank site (jackets CG3 and CB6).2.2. Acoustic Measurement EquipmentFor every measurement, a Br��el & Kj?r hydrophone (type 8104) was deployed at a depth of 10m. A Br��el & Kj?r amplifier (Nexus type 2692-0S4) was connected between the hydrophone and the recorder in order to allow for an amplification and filtration of the signal. A reference signal was used together with the output sensitivity of the Nexus to calibrate the amplitude of the recorded signal.

The signal was recorded using an audio MARANTZ Solid State Recorder (type PMD671). It was operated with the highest possible sampling rate of 44100Hz. The signal was recorded in WAVE format (.wav) on Compact Flash cards of 2GB (Sandisk Ultra II). Batteries powered all equipment. 2.3. Response VariablesIt is very common in underwater acoustics to use values expressed in a logarithmic scale (decibels). In order to characterize extreme level values of a transient signal like the one associated with pile driving the peak sound pressure level is often used. This terminology is not totally unambiguous and we prefer to use Lz?p that is defined by [9] in??dB??re??1?��Pa.(1)For impulsive sound, however,?asLz?p=10log?10pz?p2pref2 the unweighted SEL better characterises the energy Drug_discovery produced by a given stroke, extracted from a complete piling event. SEL is computed as defined by [9].

The precession of M around the total (dc and ac) magnetic field (Btot = zB0 + xBrfcos (wrft)) is given by the Bloch equations [10]:(M�BxM�ByM�Bz)=(MxMyMz)��(��Brfcos??wrft0��B0)?(��2Mx��2My��1Mz),(1)where ��1 and ��2 are the longitudinal selleck inhibitor and the transverse relaxation time, respectively. The calculated in-phase and quadrature amplitudes and the phase shift of light power with respect to the driving rf magnetic field, Brf, are given byPip(��)=?P0sin(2��)��rf�Ħ�rf2��2/��1+��22+��2;(2)Pqu(��)=?P0sin(2��)��rf��2��rf2��2/��1+��22+��2;(3)?=arctan?��2��,(4)where ��rf = �� ? wrf is the Rabi frequency, �� = wrf ? wL is the frequency detuning from the Larmor frequency, and �� is the angle between the laser beam and B0.

Several important parameters can affect the magnetometer sensitivity, namely, the laser power; the laser beam profile, the rf field power, the cell size, the buffer-gas pressure, and the density of Cs atoms (which depends on the temperature). Typically, the magnetometer spatial resolution depends on the cell dimensions. In this paper, we focus the investigation on the effects of optical power intensity and vapor cell temperature variations on the sensitivity and bandwidth of the optical Mx magnetometer.3. Experimental SetupThe Mx magnetometer used in the experiment is shown in Figures 3(a) and 3(b). The core of the instrument is a quartz-made cylindrical cell containing Cesium vapor. Also, Neon at 34Torr and Argon at 6Torr are added to the Cs vapor in order to reduce atom collisions. The cell diameter and length are 21mm and 75mm, respectively, yielding a spatial resolution of about 53mm.

In the experiments, the gas pressure inside the cell was increased by increasing the temperature using hot water flowing into a silicon pipe wrapped around the cell. The vapor cell was placed in the center of an electromagnet that generates a dc magnetic field in order to cancel the geomagnetic field and supply a uniform magnetization along the appropriate direction inside the vapor cell. The used electromagnet consists of two parts: (i) a 3D DC coils of dimension 580mm �� 530mm �� 640mm providing a magnetic field with a uniformity better than 1% in the central region and (ii) an additional pair of coils that generate a small-magnitude rf magnetic field along the x-axis.

Each coil pair of the electromagnet was independently driven by a digital power supply to cancel the geomagnetic field along the x- and y-directions and generate a uniform magnetic field along the z axis. The intensity of the magnetic field at the center of the electromagnet was 13��T, as measured by a three-axis smart digital magnetometer Honeywell HMR2300. The AC coils were driven by a waveform generator (Agilent, model 33250A) to produce an rf magnetic field of intensity 200nT, oscillating at 45.5kHz along Anacetrapib the x-axis.

The thermoelectric effect was first discovered by Thomas Seebeck in 1821 [1] when he discovered that twisting two wires together and twisting one end induced a voltage. The converse effect, that is, application CHIR-258 of voltage to induce a temperature gradient across the thermoelectric material, was discovered by Jean Peltier [1] in 1834, and is thus called the Peltier effect. The performance of the thermoelectric material is evaluated by the dimensionless figure of merit (ZT), ZT = ��S2T/K, where �� is the electrical conductivity, K is the thermal conductivity, T is the absolute temperature, and S is the Seebeck coefficient (S = ��V/��T, that is, the ratio of the induced voltage over the temperature gradient across the thermoelectric device) [2].

Thus, a high performance thermoelectric material requires high electrical conductivity, Seebeck coefficient, and low thermal conductivity. It is really challenging to find a material which has a high Seebeck coefficient (S) in combination with high electrical conductivity (��) and low thermal conductivity (k). In most materials, the electrical conductivity is directly proportional to the thermal conductivity. An ideal TE material would possess a high Seebeck coefficient as in the crystalline semiconductor, high electrical conductivity as in the crystalline metal, and low absolute temperature as in glass [4]. For practical applications, such as thermal generators (TEGs), a ZT of more than 3 is required, whilst the best efforts currently only produce a ZT of 3 [5].

Early thermoelectrical devices developed in the early 1960s earned some popularity given the solid state nature of the devices, that is, no moving parts compared to generators and motors. These devices were mainly based on Bi2Te3 [6, 7]. However, these devices have low efficiency (ZT < 1, and a system efficiency of <10%) and are therefore not cost-effective in most applications. In the mid-1990s, a research on thermoelectric started to gain interest again after theoretical predictions suggested that the thermoelectric efficiency could be enhanced through nanostructuring [8]. The introduction of nanostructures in thermoelectric materials served either to increase the electrical conductivity (through quantum dots), or to decrease the thermal conductivity (through nanowires and amorphous structures) [9�C11].

Currently, the TE communication has paid the most attention on skutterudites [12], half-Heusler alloys [13], clathrates [14], and pentafluoriade [15]. The common characteristic of these materials is their complex structure, which serves to reduce the thermal conductivity and hence increase the ZT. These materials are usually Batimastat targeted for high temperature operation, such as electricity generation from waste heat of industrial sources, such as steel furnaces and aluminum melting. This is due to their optimal ZT in a temperature range of 600K or higher.

These tubes were capped and then placed in water bath maintained at a temperature of 50��C for 48h. After this period, tubes were compound libraries taken out and cooled to room temperature. The hydrogels obtained were cut into discs of 6mm length and immersed into 50:50v/v ethanol-water solution for complete removal of catalyst and unreacted monomers. Gel discs were thoroughly washed until the pH of solution was same as the solution before washing. The hydrogels obtained were dried at 40��C until a constant weight was achieved and then were stored in vacuum desiccators for further use. Table 1Different formulations of AA/PVSA hydrogels.2.3. Swelling StudiesThe dynamic swelling ratio (Q) was evaluated gravimetrically in 100mL 0.05M USP phosphate buffer solutions of various pHs, that is, 1.2, 5.5, 6.5, and 7.

5. Preweighed dried hydrogels discs were immersed in solutions of desired pH at a temperature of 37��C. Swollen gels were removed from the medium at a predetermined interval of time for 8 hours, weighed after blotting them dry with filter paper, and placed in the same bath. The swelling ratio (Q) was calculated from the following [22]:Q=WS?WdWd,(1)where Ws and Wd are the weights of swollen and dried hydrogels, respectively.2.4. Characterization of Poly(AA-co-VSA) Hydrogels2.4.1. Structural Parameters The average molar mass of the chains between cross-links (Mc), directly related to the cross-link density, is an important parameter in characterizing the structural parameters ofhydrogels. Physical and mechanical properties of cross-linked hydrogels are significantly influenced by the Mc.

According to Flory-Rehners theory [23], Mc can be calculated usingMc=?V1dp(VS1/3?VS/2)ln?(1?VS)+VS+��VS2,(2)where VS is the volume fraction of the swollen hydrogel at equilibrium, V1 is the molar volume of the solvent (mLmol?1), �� is Flory-Huggin’s solvent interaction parameter between solvent and polymer [24], and dp is the density of gel (g mL?1).�� can be calculated from��=ln?(1?VS)+VSVS.(3)Volume fraction of the polymer (VS) was calculated by the following equation [25]:VS=[1+dpds(MaMb?1)]?1,(4)where dp and dsare densities (g mL?1) of gel and solvent, respectively. Ma and Mb are the masses (g) of the swollen and dry hydrogels, respectively.The number of links between two cross-linked chains is called cross-linked density (q) which was calculated from [26]q=dpNAMC,(5)where NA is Avogadro’s number (6.

023 �� 1023/mole).2.4.2. Fourier-Transformation Infrared Spectroscopy (FTIR) Dried discs of hydrogel samples were powdered in pestle and mortar. The powdered material was mixed with potassium bromide Drug_discovery (Merck IR spectroscopy grade) in 1:100 proportion and dried at 40��C. The mixture was compressed to semitransparent disc of 12mm diameter by applying a pressure of 65kN (pressure gauge, Shimadzu) for 2 minutes. The FTIR spectrum over the wave length range 4000�C400cm?1 was recorded using FTIR spectrometer (FTIR 8400 S, Shimadzu).2.4.3.

Ultrapure water Volasertib cancer (resistance > 18m��) was obtained from Milli-Q system (Millipore, France). 2.2. Standard SolutionsStock standard solutions (1mg/mL) were prepared by weighing 25.0mg of each reference standard and dissolving in 25mL methanol. These solutions were kept in the temperature below ?18��C for 12 months. Mixed working standard solution (20��g/mL MAD and 100��g/mL of remaining ionophores) was stored in 6�C10��C up to three months.2.3. Sample TreatmentFor the method development and validation, animal feed samples without ionophore coccidiostats were used. The feed was ground with rotor mill ZM200 (Retsch, Germany) and sieved through 0.5mm sieve. The sample (5g �� 0.01g) was weighted into the polypropylene centrifuge tube.

The appropriate amount of mixed standard solution was added to the spiked samples, and the sample was let to stand for at least one hour. Methanol was added to the sample so that the total solvent volume was 25mL (the volume of methanol added with the standard solution was subtracted), and the sample was shaken for 30 minutes at 200 cycles/min (MaxQ 2000 Orbital Shaker, Thermo Scientific, USA). The sample was then centrifuged (3500rpm, 10min), and 0.5mL of supernatant was transferred to a glass tube. Internal standard (monensin methyl ester, 10��L of 10��g/mL solution) and 0.5mL DMSO were added, and the sample was evaporated under the stream of nitrogen (45��C). The extract in DMSO was transferred into a vial and analysed with liquid chromatography.2.4.

Instrumental AnalysisThe instrumental analysis of coccidiostats was performed using Varian Prostar HPLC system equipped with quaternary pump, autosampler, column oven, postcolumn derivatisation module, and two detectors��fluorescence and UV-Vis, controlled by Galaxie Workstation software. Chromatographic separation of compounds was performed on Kinetex C18 column (150 �� 4.6mm, 2.6��m, Phenomenex, USA) connected with precolumn (4 �� 3mm, SecurityGuard, Phenomenex, USA). The isocratic elution was applied, with mobile phase consisting of 88% methanol and 12% 0.02M KH2PO4, adjusted to pH 7.0 Anacetrapib at 0.7mL/min flow rate. Column oven temperature was controlled at 32��C. After HPLC separation, eluate was transferred through fluorescence detector (excitation and emission wavelength 310 and 420nm, resp.) for the lasalocid detection. Next, the eluate was passed through the reaction cell (1.4mL) of the postcolumn derivatisation reactor (Pinnacle PCX, Pickering, USA). The derivatisation was performed with 6% vanillin solution in methanol (0.35mL/min) in the presence of 4% sulfuric acid in methanol (0.35mL/min), and the coil was maintained at 110��C. The derivatives of ionophores were then detected at �� 520nm. The injection volume was 50��L.2.5. Validation2.5.1.

Gu et al. (2012). This study explored the post-ischemic function of all subsets of T cells, aiming at clarifying the protective or detrimental roles of distinctive subsets after stroke. In this study, CD8+ cytotoxic T cells, CD4+ Th1 cells, CD4+ Th2 cells, and Tregs were evaluated both in vivo and in vitro, by means etc of genetic knockout of each subset and coculture of neurons with splenocytes from each knockout. As was illustrated in this experiment, elimination of Treg did not influence the outcome, yet deficiency of Th1 would attenuate while Th2 knockout would aggravate inflammatory response in vivo and neuronal death in vitro.Stubbe et al. (2013). This study was mainly focused on the accumulation, proliferation, and function of endogenous Tregs in a late phase after stroke.

By means of fluorescence activated cell sorting analysis and immunohistochemistry, the authors demonstrated that after MCAO, Tregs began to accumulate in the infarct and peri-infarct areas on day 7, and the ipsilesional accumulation persisted till day 30. Treg proliferation in the infarct areas increased throughout days 7 and 14 before its declination to a nearly normal level by day 30. Moreover, in order to clarify the late function of Tregs, 30min MCAO was performed followed by anti-CD25 mAb administered on days 3 and 14. Infarct volume was measured by MRI on days 3 and 27, and neural function was evaluated by gait analysis on days 14 and 27, but no significant difference was observed between the delayed Treg depletion group and controls.4. Discrepancies and DiscussionAs mentioned above, Tregs yield to profound inconstancy.

The controversial topic has triggered intensified debate and discussion recently. Outcome of current studies in murine model was summarized in Table 1. Table 1Outcome of current studies in murine model.4.1. Whether Endogenous Tregs Depletion Benefits or Exacerbates the OutcomeOne of the major discrepancies posed by Liesz et al. [55] and Kleinschnitz et al. [50] is whether endogenous Tregs are relevant to secondary infarct growth after stroke. Liesz et al. proved that Treg depletion was associated with secondary infarct growth by day 7, although these effects would not become evident within the first 3 days, which was consistent with the neutral report from Ren et al. [60] and Gu et al. [61], whereas in the experiment conducted by Kleinschnitz et al.

, Treg depletion led to a better outcome within 24 hours and Entinostat no progression occurred in both groups out to 1 week.A plausible explanation for the inconsistency may be attributed to the ischemic duration and measured time because specific cell types of the immune system might be differentially relevant in different models of stroke [52]. The most part of the study conducted by Liesz et al. was based on a permanent MCAO mouse model inducing cortical infarction around 15mm3 in size, while in the experiment reported by Kleinschnitz et al.

Accordingly, the direction of outflow arterial flow and distance from coronaries is also of importance [28]. Our results add to the controversy about the effect of IABP-generated pulsatility both on macro- and microcirculation; in small non-randomized clinical apply for it studies, Jung [36,37] and den Uil [38] have reported on a favorable effect of IABP on microcirculation in cardiogenic shock, but Mustermann [39] showed paradoxically improved microvascular flow after withdrawing the IABP. Microcirculation changes definitely play a key role in critical states [40-42] and the role of pulsatility and different support combinations (that is, central vs. peripheral, subclavian vs. femoral) on microcirculatory changes remains to be further studied.

An important observation of our study is that VA ECMO significantly increases coronary perfusion pressure over time, mainly in the FF configuration. CoPP actually reached the baseline values by the end of our protocol, that is, approximately two hours after the ECMO onset (Figures (Figures33 and and4,4, Additional file 4 and Table Table3).3). When pooling data from all animals, the CoPP increased progressively from 15.0 mmHg at the end of the cardiac arrest to 34.0 mmHg five minutes after starting ECMO, rose to 53.0 mmHg at the ECMO switch and continued to rise to 68.0 mmHg before CPR, not different from baseline. The increases in CoPP were more pronounced during the FF ECMO period of the protocol, which was responsible more than FS ECMO for an overall CoPP increase.

In contrast to FF ECMO, where baseline to before CPR values reached the same values, in the FS ECMO starting cohort, the CoPP at the end of the protocol still remained significantly lower compared to baseline (45.5 vs. 78.5 mmHg, respectively, P = 0.041). To our knowledge, this phenomenon of the CoPP increase over time during VA ECMO treated cardiac arrest has not been described before. This finding is important, because CoPP is a key prerequisite for ROSC in prolonged cardiac arrest [43,44]. We can only speculate about the pathophysiological mechanism behind the low CoPP during ECMO start and gradual CoPP increase with time on ECMO; however, vasoplegia with low peripheral resistance post low flow cardiac arrest followed by improved vasoreactivity during further ECMO reperfusion offers a reasonable explanation.

We intentionally did not use vasopressors in this phase of the experiment in order to observe the ?natural” course of ECMO reperfusion. In human care, we are used to keeping the perfusion pressure (that is, MAP) within optimal range with norepinephrine to assure adequately both cerebral flow and perfusion pressure.The adequacy of CoPP was reflected by a high resuscitability of our animals (see Additional file 5), despite a rather Cilengitide strenuous and prolonged protocol.

Then, the conidia from the nonselective plates were plated-out on CD selective medium containing 5-FOA, uracil and uridine. This led to isolation of the mutant strain, GR6 kinase inhibitor Ruxolitinib pyrG��0. PCR based tests confirmed that the pyrG region of strain GR6 was deleted.It was possible to complement the uridine requirement of GR6 pyrG��0 by introducing the plasmids pUC_pyrGAo and ANEp2 carrying A. oryzae or A. nidulans pyrG, respectively. This suggests that the AMA1 region of ANEp2, which supports autonomous plasmid replication in other fungi, also supports autonomous replication in A. oryzae. Furthermore, the transformation frequency of ANEp2 was about 10-fold greater than that obtained with the integrative plasmid pUC_pyrGAo.

Transformation frequency ratios obtained with pUC_pyrGAo as compared to plasmids harbouring AMA1 were in accordance with those obtained with integrating versus autonomously replicating plasmids [12]. Also, Fierro et al. [21] transformed P. nalgiovense with integrative versus autoreplicative plasmids and found 60-fold higher frequencies for AMA1-based autonomously replicating plasmids compared with integrative plasmids. Autonomously replicating plasmids carrying the AMA1 region from A. nidulans transformed P. nalgiovense very efficiently and these plasmids also showed mitotic stability with low degrees of reorganization.��-Galactosidase expression showed that the A. oryzae strain GR6 pyrG��0 was able to efficiently produce heterologous secreted proteins using the A. niger transcriptional and translational controls introduced via ANEp2.

The strain GR6 pyrG��0 transformed with ANEp2 containing lacA encoding A. niger ��-galactosidase was more than 150 times higher than that obtained with the untransformed strain. The finding of this study is promising and this paves the way for the application of A. oryzae strain GR6 pyrG��0 in heterologous protein production given that this strain is amenable to genetic manipulation and that A. oryzae is widely used in biotechnology and the food industry.5. ConclusionMutant strain GR6, a pyrG��0 derivative of the A. oryzae strain S1, was constructed using targeted gene replacement. The pyrG auxotrophs were verified by assessing their ability to grow on selective and nonselective media and PCR analysis. Plasmids pUC_pyrGAo and ANEp2, harbouring the pyrG genes from A. oryzae and A.

nidulans, respectively, complemented the uracil and uridine requirement of strain GR6. Transformants of the A. oryzae GR6 mutant carrying the heterologous expression vector ANEp2 were able to functionally express a secreted ��-galactosidase at high levels.Supplementary MaterialThe cassette used for deleting the Drug_discovery pyrG gene was 3,568bp long. Boxed and lowercase letters indicated the XhoI site which fused the pyrG 5�� and 3�� UTR, respectively.