Efforts to develop sustainable industrial processes have led to significant advances toward supplanting petrochemical‐dependent technologies. Some of these otherwise sustainable processes, notably animal product rendering and biodiesel production, produce low value waste that is high in free fatty acids. Sulfur in turn is a primary waste product of fossil fuel refining. In the current contribution, copolymers are prepared by reaction of elemental sulfur with fatty acids in several monomer ratios. Both monounsaturated oleic acid and bis(unsaturated) linoleic acid were evaluated to assess the extent to which copolymer properties relate to the degree of unsaturation of the fatty acid comonomer. Furthermore, copolymers prepared from technical grade versus pure linoleic acid were compared to evaluate the viability of the considerably more affordable technical grade monomer. The thermal and mechanical properties of the copolymers were assessed by thermogravimetric analysis, differential scanning calorimetry and dynamic mechanical analysis. © 2020 Wiley Periodicals, Inc. J. Polym. Sci.
- Award ID(s):
- 1827209
- PAR ID:
- 10174163
- Date Published:
- Journal Name:
- 2019 ACS Midwest Regional Meeting
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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ABSTRACT 2020 ,58 , 438–445 -
Slotman, Michel (Ed.)Abstract The wide distribution of Culex (Cx.) pipiens complex mosquitoes makes it difficult to prevent the transmission of mosquito-borne diseases in humans. Gene editing using CRISPR/Cas9 is an effective technique with the potential to solve the growing problem of mosquito-borne diseases. This study uses the ReMOT Control technique in Culex pipiens pallens (L.) to produce genetically modified mosquitoes. A microinjection system was established by injecting 60 adult female mosquitoes—14 µl injection mixture was required, and no precipitation occurred with ≤1 µl of endosomal release reagents (chloroquine or saponin). The efficiency of delivery of the P2C-enhanced green fluorescent protein-Cas9 (P2C-EGFP-Cas9) ribonucleoprotein complex into the ovary was 100% when injected at 24 h post-bloodmeal (the peak of vitellogenesis). Using this method for KMO knockout, we found that gene editing in the ovary could also occur when P2C-Cas9 RNP complex was injected into the hemolymph of adult Cx. pipiens pallens by ReMOT Control. In the chloroquine group, of the 2,251 G0 progeny screened, 9 individuals showed with white and mosaic eye phenotypes. In the saponin group, of the 2,462 G0 progeny screened, 8 mutant individuals were observed. Sequencing results showed 13 bp deletions, further confirming the fact that gene editing occurred. In conclusion, the successful application of ReMOT Control in Cx. pipiens pallens not only provides the basic parameters (injection parameters and injection time) for this method but also facilitates the study of mosquito biology and control.more » « less
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Summary Volvox carteri and other volvocine green algae comprise an excellent model for investigating developmental complexity and its origins. Here we describe a method for targeted mutagenesis inV. carteri usingCRISPR /Cas9 components expressed from transgenes. We usedV. carteri nitrate reductase gene (nitA ) regulatory sequences to conditionally expressStreptococcus pyogenes Cas9, andV. carteri U6RNA gene regulatory sequences to constitutively express single‐guideRNA (sgRNA ) transcripts.Volvox carteri was bombarded with both Cas9 vector and one of several sgRNA vectors programmed to target different test genes (glsA ,regA andinvA ), and transformants were selected for expression of a hygromycin‐resistance marker present on the sgRNA vector. Hygromycin‐resistant transformants grown with nitrate as sole nitrogen source (inducing fornitA ) were tested for Cas9 and sgRNA expression, and for the ability to generate progeny with expected mutant phenotypes. Some transformants of a somatic regenerator (Reg) mutant strain receiving sgRNA plasmid withglsA protospacer sequence yielded progeny (at a rate of ~0.01%) with a gonidialess (Gls) phenotype similar to that observed for previously describedglsA mutants, and sequencing of theglsA gene in independent mutants revealed short deletions within the targeted region ofglsA , indicative of Cas9‐directed non‐homologous end joining. Similarly, bombardment of a morphologically wild‐type strain with the Cas9 plasmid and sgRNA plasmids targetingregA orinvA yieldedregA andinvA mutant transformants/progeny, respectively (at rates of 0.1–100%). The capacity to make precisely directed frameshift mutations should greatly accelerate the molecular genetic analysis of development inV. carteri , and of developmental novelty in the volvocine algae. -
The DNA inside human cells provides instructions for all of the processes that happen inside the body. Errors in the DNA may lead to cancer, sickle cell disease, cystic fibrosis, Huntington’s disease, or other genetic disorders. Medical researchers are exploring whether it is possible to replace or repair the faulty DNA (an approach known as gene therapy) to reduce the symptoms, or even cure individuals, of these conditions. Over the last ten years, a new technology known as CRISPR-Cas9 gene editing has proved to be a reliable and efficient way to make small and precise changes to DNA in living cells. First, an enzyme called Cas9 searches for a segment of target DNA segment that matches a template molecule the enzyme carries. Cas9 then cuts the target DNA, which is repaired to match a new customized DNA sequence: this changes the genetic information of the cell. The Cas9 protein is made of a succession of building blocks called amino acids that create long chains which then fold to form the final three-dimensional shape of the enzyme. A region of Cas9 known as the HNH domain is responsible for cutting the target DNA. However, it remains unclear exactly which amino acids within this domain work together to sever the DNA. Here, Zuo et al. combined computational and experimental approaches to reveal the three-dimensional structure of the Cas9 enzyme when the HNH domain is poised to cut the target DNA. The findings were used to generate a computational model of Cas9 and this model predicted that the HNH domain relies on a group of three amino acids known collectively as D839-H840-N863 to cleave DNA strands. This knowledge is useful to understand exactly how Cas9 modifies genetic information. Ultimately, this may help to improve CRISPR-Cas9 technology so it could be safely used in geneediting therapies.more » « less
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Abstract CRISPR‐Cas9 has been shown to be a valuable tool in recent years, allowing researchers to precisely edit the genome using an RNA‐guided nuclease to initiate double‐strand breaks. Until recently, classical RAD51‐mediated homologous recombination has been a powerful tool for gene targeting in the moss
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