Synthesis of 3-substituted 2-cyclohexenones through umpoled functionalization

El trabajo se refiere a una metodología sobre un estudio de reactividad invertidaA new protocol to obtain 3-substituted 2-cyclohexenones, was developed by reversing the chemical reactivity of 2-cyclohexenone. One-pot synthesis of 3-substituted 2-cyclohexenones can be achieved by treatment of 3-phenylthiosilyl enol ether with a mixture of t-BuLi/HMPA that allows hydrogen-selective exchange in presence of reactive electrophiles such as aldehydes, ketones and alkyl halides. This affords the corresponding product in moderate overall yield, after silyl enol ether cleavage and concomitant thiophenol elimination initiated with TBAF.Conacy

Toward the definition of efficacy and safety criteria for advancing gene drive-modified mosquitoes to field testing

Mosquitoes containing gene drive systems are being developed as complementary tools to prevent transmission of malaria and other mosquito-borne diseases. As with any new tool, decision makers and other stakeholders will need to balance risks (safety) and benefits (efficacy) when considering the rationale for testing and deploying gene drive-modified mosquito products. Developers will benefit from standards for judging whether an investigational gene drive product meets acceptability criteria for advancing to field trials. Such standards may be formalized as preferred product characteristics and target product profiles, which describe the desired attributes of the product category and of a particular product, respectively. This report summarizes discussions from two scientific workshops aimed at identifying efficacy and safety characteristics that must be minimally met for an investigational gene drive-modified mosquito product to be deemed viable to move from contained testing to field release and the data that will be needed to support an application for first field release

A CRISPR–Cas9 gene drive targeting doublesex causes complete population suppression in caged Anopheles gambiae mosquitoes

In the human malaria vector Anopheles gambiae, the gene doublesex (Agdsx) encodes two alternatively spliced transcripts, dsx-female (AgdsxF) and dsx-male (AgdsxM), that control differentiation of the two sexes. The female transcript, unlike the male, contains an exon (exon 5) whose sequence is highly conserved in all Anopheles mosquitoes so far analyzed. We found that CRISPR-Cas9-targeted disruption of the intron 4-exon 5 boundary aimed at blocking the formation of functional AgdsxF did not affect male development or fertility, whereas females homozygous for the disrupted allele showed an intersex phenotype and complete sterility. A CRISPR-Cas9 gene drive construct targeting this same sequence spread rapidly in caged mosquitoes, reaching 100% prevalence within 7-11 generations while progressively reducing egg production to the point of total population collapse. Owing to functional constraint of the target sequence, no selection of alleles resistant to the gene drive occurred in these laboratory experiments. Cas9-resistant variants arose in each generation at the target site but did not block the spread of the drive

Immobilised teicoplanin does not demonstrate antimicrobial activity against Staphylococcus aureus

Abstract: Antibacterial bone biomaterial coatings appeal to orthopaedics, dentistry and veterinary medicine. Achieving the successful, stable conjugation of suitable compounds to biomaterial surfaces is a major challenge. A pragmatic starting point is to make use of existing, approved antibiotics which are known to remain functional in a stationary, immobilised state. This includes the macrocyclic glycopeptide, teicoplanin, following the discovery, in the 1990’s, that it could be used as a chiral selector in chromatographic enantiomeric separations. Importantly teicoplanin works at the level of the bacterial cell wall making it a potential candidate for biomaterial functionalisations. We initially sought to functionalise titanium (Ti) with polydopamine and use this platform to capture teicoplanin, however we were unable to avoid the natural affinity of the antibiotic to the oxide surface of the metal. Whilst the interaction between teicoplanin and Ti was robust, we found that phosphate resulted in antibiotic loss. Before contemplating the covalent attachment of teicoplanin to Ti we examined whether a commercial teicoplanin stationary phase could kill staphylococci. Whilst this commercially available material could bind N-Acetyl-L-Lys-D-Ala-D-Ala it was unable to kill bacteria. We therefore strongly discourage attempts at covalently immobilising teicoplanin and/or other glycopeptide antibiotics in the pursuit of novel antibacterial bone biomaterials

Board of Directors Meeting Minutes (May 8, 2006)

This file contains the minutes from the Des Moines Area Community College Board meeting held on May 8, 2006

Behavior of homing endonuclease gene drives targeting genes required for viability or female fertility with multiplexed guide RNAs

A gene drive method of particular interest for population suppression utilizes homing endonuclease genes (HEGs), wherein a site-specific, nuclease-encoding cassette is copied, in the germline, into a target gene whose loss of function results in loss of viability or fertility in homozygous, but not heterozygous, progeny. Earlier work in Drosophila and mosquitoes utilized HEGs consisting of Cas9 and a single guide RNA (gRNA) that together target a specific gene for cleavage. Homing was observed, but resistant alleles immune to cleavage, while retaining wild-type gene function, were also created through nonhomologous end joining. Such alleles prevent drive and population suppression. Targeting a gene for cleavage at multiple positions has been suggested as a strategy to prevent the appearance of resistant alleles. To test this hypothesis, we generated two suppression HEGs in Drosophila melanogaster targeting genes required for embryonic viability or fertility, using a HEG consisting of CRISPR/Cas9 and gRNAs designed to cleave each gene at four positions. Rates of target locus cleavage were very high, and multiplexing of gRNAs prevented resistant allele formation. However, germline homing rates were modest, and the HEG cassette was unstable during homing events, resulting in frequent partial copying of HEGs that lacked gRNAs, a dominant marker gene, or Cas9. Finally, in drive experiments, the HEGs failed to spread due to the high fitness load induced in offspring as a result of maternal carryover of Cas9/gRNA complex activity. Alternative design principles are proposed that may mitigate these problems in future gene drive engineering

Determining the landscape of resistance to gene drives in the malaria mosquito

Gene drives are engineered selfish genetic elements with the potential to spread throughout entire insect populations for sustainable vector control. Recently, a gene drive was shown to eliminate caged populations of the malaria mosquito by targeting the highly conserved female-specific exon of the doublesex gene. This caused females, homozygous for the gene drive, to develop as sterile intersex individuals, leading to the observed population crash. However, target site resistant alleles that block gene drive activity, whilst encoding a functional copy of the target gene, may halt gene drive spread in the wild. These may be naturally occurring or generated by the gene drive itself. This thesis presents a pipeline for the discovery, genetic engineering, and testing of putative drive-resistant variants. First, to investigate the potential for natural resistance, existing population genomics data were interrogated for the presence of natural single nucleotide polymorphisms (SNPs) at the highly conserved gene drive target region. To investigate the potential for drive-induced resistance, a high-throughput assay was designed to generate a high volume of mutations at the gene drive target site and screen them for their ability to restore dsx function. These methods yielded three putatively resistant SNPs: one natural polymorphism and two rare Cas9-induced mutations. These were engineered in the mosquito genome for testing, using a novel method termed CRISPR-mediated cassette exchange (CriMCE). It was confirmed that all three polymorphisms are functional and offer full, partial or no resistance to gene drive. Importantly, partial resistance to gene drive is being demonstrated for the first time. To mitigate observed resistance, gene drive systems targeting multiple sites simultaneously were developed. These showed improved drive dynamics and caused rapid elimination of caged mosquito populations within 7-8 generations. The experimental pipeline described here can be applied to pre-empt and mitigate resistance against any gene drive strategy, prior to field testing.Open Acces

Työntekijöiden kokemuksia ammatillisesta kehittymisestä etätyössä

Tämän tutkielman tavoitteena oli tutkia ammatillisen kehittymisen sekä sosiaalisen kanssakäymisen välistä suhdetta etätyön kontekstissa kohdeorganisaatiossa, joka on henkilöstöhallinnon tukipalveluita tuottava asiantuntijayksikkö. Etätyön tutkimuksissa, fyysisellä etäisyydellä on todettu olevan negatiivinen vaikutus työntekijöiden välisten suhteiden laatuun, joka saattaa heikentää tiedon sekä osaamisen jakamista työyhteisössä, ja siten vaikuttaa ammatilliseen kehittymiseen heikentävästi. Ammatillista kehittymistä tarkasteltiin tässä tutkielmassa erityisesti sosiaalisen pääoman näkökulmasta, joka voidaan yhdistää sosiaaliseen kanssakäymiseen, sillä sosiaalisella pääomalla on tutkimusten mukaan todettu olevan positiivinen vaikutus tiedon sekä osaamisen jakamiseen organisaatioissa ja siten myös positiivinen vaikutus yksilöiden, tiimien sekä organisaatioiden osaamisen kehittämiselle. Työntekijän oma tulkinta katsottiin tutkimuksen kannalta oleelliseksi, sillä työntekijän oma tulkinta ja kokemus omista kyvyistään sekä osaamisestaan voidaan katsoa merkittäväksi tekijäksi työssä toimimiselle. Työntekijän pystyvyysuskomuksilla on myös vaikutusta aktiivisuuteen ammatillista kehittämistä kohtaan haastavampien tehtävien kautta. Fenomenologis-hermeneuttisen taustafilosofian mukaisesti tässä tutkimuksessa pyrit-tiin sekä ymmärtämään työntekijöiden kokemuksia niin kuin työntekijät ne ymmärtävät sekä tuoda näitä kokemuksia esiin tulkinnan kautta. Tuloksia tulkittiin seitsemän työntekijän kokemuksien kautta. Tutkimusaineiston perusteella työntekijät kokivat sosiaalisen pääoman heikentyneen etätyöaikana, joka näkyi yhteistyön ja yhteisöllisyyden vähenemisenä sekä tiedon sekä osaamisen jakamisen haasteina vaikuttaen myös ammatilliseen kehittymiseen heikentävästi. Etätyössä työtovereiden merkitys työtehtävissä selviytymiselle sekä työhyvinvoinnille korostui entisestään. Esimiehen sekä työntekijöiden väliseen suhteeseen vaikuttaa sekä kommunikoinnin tiheys että sen laatu, jotka korostuvat etätyössä. Esimiehen palautteella, tuella sekä työtehtävien kautta luoduilla mahdollisuuksilla koettiin olevan ammatillista kehittymistä edistävä vaikutus. Ammatillista kehittymistä etätyössä edistää myös työntekijän oma aktiivisuus sekä sosiaalisen verkostoon osallistuminen tietoa sekä osaamista jakamalla