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The Adapting Agriculture to Climate Change Project set out to improve the diversity, quantity, and accessibility of germplasm collections of crop wild relatives (CWR). Between 2013 and 2018, partners in 25 countries, heirs to the globetrotting legacy of Nikolai Vavilov, undertook seed collecting expeditions targeting CWR of 28 crops of global significance for agriculture. Here, we describe the implementation of the 25 national collecting programs and present the key results. A total of 4587 unique seed samples from at least 355 CWR taxa were collected, conserved ex situ, safety duplicated in national and international genebanks, and made available through the Multilateral System (MLS) of the International Treaty on Plant Genetic Resources for Food and Agriculture (Plant Treaty). Collections of CWR were made for all 28 targeted crops. Potato and eggplant were the most collected genepools, although the greatest number of primary genepool collections were made for rice. Overall, alfalfa, Bambara groundnut, grass pea and wheat were the genepools for which targets were best achieved. Several of the newly collected samples have already been used in pre-breeding programs to adapt crops to future challenges.

In this work we hypothesized that the chemokine fractalkine can serve as a cancer molecular target. We engineered aptamer micelles functionalized with an outer poly(ethylene glycol) (PEG) corona, and investigated the extent and efficacy of using them as a targeting tool against fractalkine-expressing colon adenocarcinoma cells. In vitro cell binding results showed that aptamer micelles bound and internalized to fractalkine-expressing cancer cells with the majority of the micelles found free in the cytoplasm. Minimal surface binding was observed by healthy cells. Even though partial PEGylation did not prevent serum adsorption, micelles were highly resistant to endonuclease and exonuclease degradation. In vivo biodistribution studies and confocal studies demonstrated that even though both aptamer and control micelles showed tumor accumulation, only the aptamer micelles internalized into fractalkine-expressing cancer cells, thus demonstrating the potential of the approach and showing that fractalkine may serve as a specific target for nanoparticle delivery to cancer cells., (Copyright © 2017 Elsevier Inc. All rights reserved.)

A set of poly(δ-valerolactone- co -d,l-lactide)- b -poly(ethylene glycol)- b -poly(δ-valerolactone- co -d,l-lactide) (PVLA-PEG-PVLA) triblock copolymers was synthesized and the solution properties were characterized using rheology, cryo-TEM, cryo-SEM, SANS, and degradation studies. This polymer self-assembles into a low viscosity fluid with flowerlike spherical micelles in water at room temperature and transforms into a wormlike morphology upon heating, accompanied by gelation. At even higher temperatures syneresis is observed. At physiological temperature (37 °C) the hydrogel's average pore size is around 600 nm. The PVLA-PEG-PVLA gel degrades in about 45 days in cell media, making this unique hydrogel a promising candidate for biomedical applications.

Ex situ conservation efforts such as those of zoos, botanical gardens, and seed banks will form a vital complement to in situ conservation actions over the coming decades. It is therefore necessary to pay the same attention to the biological diversity represented in ex situ conservation facilities as is often paid to protected-area networks. Building the phylogenetic diversity of ex situ collections will strengthen our capacity to respond to biodiversity loss. Since 2000, the Millennium Seed Bank Partnership has banked seed from 14% of the world's plant species. We assessed the taxonomic, geographic, and phylogenetic diversity of the Millennium Seed Bank collection of legumes (Leguminosae). We compared the collection with all known legume genera, their known geographic range (at country and regional levels), and a genus-level phylogeny of the legume family constructed for this study. Over half the phylogenetic diversity of legumes at the genus level was represented in the Millennium Seed Bank. However, pragmatic prioritization of species of economic importance and endangerment has led to the banking of a less-than-optimal phylogenetic diversity and prioritization of range-restricted species risks an underdispersed collection. The current state of the phylogenetic diversity of legumes in the Millennium Seed Bank could be substantially improved through the strategic banking of relatively few additional taxa. Our method draws on tools that are widely applied to in situ conservation planning, and it can be used to evaluate and improve the phylogenetic diversity of ex situ collections., (© 2014 Society for Conservation Biology.)

DNA nanotubes were created using molecular self-assembly of single-stranded DNA (ssDNA)-amphiphiles composed of a hydrophobic dialkyl tail and polycarbon spacer and a hydrophilic ssDNA headgroup. The nanotube structures were formed by bilayers of amphiphiles, with the hydrophobic components forming an inner layer that was shielded from the aqueous solvent by an outer layer of ssDNA. The nanotubes appeared to form via an assembly process that included transitions from twisted nanotapes to helical nanotapes to nanotubes. Amphiphiles that contained different ssDNA headgroups were created to explore the effect of the length and secondary structure of the ssDNA headgroup on the self-assembly behavior of the amphiphiles in the presence and absence of the polycarbon spacer. It was found that nanotubes could be formed using a variety of headgroup lengths and sequences. The ability to create nanotubes via ssDNA-amphiphile self-assembly offers an alternative to the other purely DNA-based approaches like DNA origami and DNA tile assembly for constructing these structures and may be useful for applications in drug delivery, biosensing, and electronics.

Previously we identified an aptamer, named FKN-S2, which binds the cell surface protein fractalkine with high affinity and specificity. In this paper a hydrophobic dialkyl C16 tail was added to the aptamer to create an aptamer-amphiphile. We investigated how the tail and a spacer molecule of varying length and hydrophobicity, inserted between the tail and the aptamer headgroup, affect the binding, structure, and self-assembly properties of the aptamer-amphiphile. We synthesized aptamer-amphiphiles with no spacer (NoSPR), polyethylene glycol (PEG4, PEG8, PEG24), alkyl (C12 and C24), or oligonucleotide (T10 and T5: 10 and 5 thymine, and A10: 10 adenine) spacers. The addition of the tail reduced the binding affinity of the aptamer-amphiphile over 7.5-fold compared to the free aptamer. The hydrophobic alkyl spacers resulted in the greatest loss of affinity, and the hydrophilic PEG spacers improved amphiphile affinity but did not restore it to that of the free aptamer. Interestingly, oligonucleotide spacers produced the highest affinity amphiphiles. Nucleotide composition did not affect affinity, however, as the T10 and A10 spacers had equal affinity. The oligonucleotide spacer amphiphiles had the highest affinity because the oligonucleotide spacer increased the affinity of free aptamer; the FKN-S2 aptamer plus the oligonucleotide spacer had a higher affinity than the free FKN-S2 aptamer. Circular dichroism (CD) spectroscopy and thermal melting studies indicated the aptamer forms a stem-loop and intramolecular G-quadruplex, and the tail strongly stabilized the formation of the G-quadruplex in a buffer. Cryogenic transmission electron microscopy (cryo-TEM) imaging showed the aptamer-amphiphiles, independent of the spacer used, self-assembled into micelles and nanotapes, flat bilayer structures that were often twisted. Finally, liposomes functionalized with the FKN-S2 amphiphile were incubated with fractalkine expressing cells, and the amount of binding was dependent on the concentration of the amphiphile on the liposome surface.

The self-assembly of single-stranded DNA (ssDNA) aptamer-amphiphiles was influenced by the choice of spacer used to link the hydrophobic tail and aptamer headgroup. Aptamer-amphiphiles without spacers or with hydrophilic spacers formed globular micelles while amphiphiles with hydrophobic spacers formed bilayer nanotapes, which are the first such structures formed by DNA-amphiphiles.

The success of common nonviral gene delivery vehicles, lipoplexes and polyplexes, is limited by the toxicity and instability of these charged molecules. Stealth liposomes could provide a stable, safe alternative to cationic DNA complexes for effective gene delivery. DNA encapsulations in three stealth liposomal formulations prepared by thin film, reverse phase evaporation, and asymmetric liposome formation were compared, and the thin film method was found to produce the highest yields of encapsulated DNA. A DNA quantification method appropriate for DNA encapsulated within liposomes was also developed and verified for accuracy. The effect of initial lipid and DNA concentrations on the encapsulation yield and fraction of DNA-filled liposomes was evaluated. Higher encapsulation yields were achieved by higher lipid contents, while a higher fraction of DNA-filled liposomes was produced by either lower lipid content or higher DNA concentration. Control of these parameters allows for the design of gene delivery nanoparticles with high DNA encapsulation yields or higher fraction of DNA-filled liposomes.

A gene delivery system was designed to carry a payload to integrin overexpressing cells. Branched-polyethyleneimine (bPEI) condensed plasmid DNA was encapsulated into targeted stealth liposomes, thereby combining the condensing and transfection properties of bPEI with the stealth and targeting properties of the liposomal carrier system. PR_b was used as a targeting ligand - a peptide we designed to bind specifically to the cancer cell surface marker α 5 β 1 integrin - and such a robust receptor-ligand interaction achieved higher specificity than what has been previously reported for targeted delivery systems. In the process of formulating the PR_b functionalized gene delivery vehicle, we developed a protocol to fully encapsulate condensed DNA in liposomes and accurately quantify the total DNA in the system. We demonstrate that compared to non-targeted stealth liposomes and non-encapsulated condensed DNA, the PR_b functionalized stealth liposomes mediated improved in vitro transfection specifically to colon cancer cells overexpressing the α 5 β 1 integrin. Furthermore, when administered in vivo to metastatic tumor bearing mice, PR_b functionalized stealth liposomes outperformed non-targeted liposomes and delivered genes specifically to the tumor site.

Fractalkine (FKN) is a unique cell surface protein with potential as a therapeutic target because of its role in inflammatory diseases and cancer. We developed an aptamer, named FKN-S2, with a dissociation constant of 3.4 ± 0.7 nM that is specific to the chemokine domain of fractalkine.

Encapsulating anticancer drugs in nanoparticles has proven to be an effective mechanism to alter the pharmacokinetic and pharmacodynamic profiles of the drugs, leading to clinically useful cancer therapeutics like Doxil and DaunoXome. Underdeveloped tumor vasculature and lymphatics allow these first-generation nanoparticles to passively accumulate within the tumor, but work to create the next-generation nanoparticles that actively participate in the tumor targeting process is underway. Lipid nanoparticles functionalized with targeting peptides are among the most often studied. The goal of this article is to review the recently published literature of targeted nanoparticles to highlight successful designs that improved in vivo tumor therapy, and to discuss the current challenges of designing these nanoparticles for effective in vivo performance., (Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

In recent years, a variety of biomimetic constructs have emerged which mimic the bioactive sequences found in the natural extracellular matrix (ECM) proteins such as fibronectin (FN) that promote cell adhesion as well as proliferation on artificially functionalized interfaces. Much interest lies in investigating the ability of the ECM mimetic materials in regulating a number of vital cell functions including differentiation, gene expression, migration, and proliferation. A peptide amphiphile PR_b containing both the cell adhesive GRGDSP and synergistic PHSRN peptide sequences was developed in our group that was shown to support enhanced cell proliferation and ECM FN secretion as compared to GRGDSP and FN functionalized interfaces. In this study, we have investigated the binding affinity of the PR_b peptide ligand with the FN cell surface receptor, the α(5)β(1) integrin. We compared PR_b functionalized surfaces with FN and BSA coated surfaces and GRGDSP functionalized surfaces in terms of promoting intracellular signaling cascades that are essential for enhanced cellular activity. Specifically, we studied the phosphorylation of focal adhesion kinase (FAK) at tyrosine residues Y397 and Y576 and the formation of cyclin D1, both of which are intracellular markers of integrin mediated attachment of cells, signaling pathways, and progression of cell cycle. FAK and cyclin D1 encourage enhanced cell proliferation, differentiation, and gene expression. Our results show that the PR_b peptide ligand has a specific and strong binding affinity for the α(5)β(1) integrin with a dissociation constant of 76.3 ± 6.3 nM. The PR_b peptide ligands supported enhanced FAK phosphorylation activity and increased cyclin D1 formation as compared to the widely used GRGDSP ligand, the native protein FN (positive control), and BSA nonadhesive surfaces (negative control). These results encourage the use of the FN mimetic PR_b peptide in functionalizing biomaterials for potential tissue engineering and therapeutic applications.

To analyze the spatiotemporal dynamics of network activity in a brain tissue slice, it is useful to record simultaneously from multiple locations. When obtained from laminar structures such as the hippocampus or neocortex, multisite recordings also yield information about subcellular current distributions via current source density analysis. Multisite probes developed for in vivo recordings could serve these purposes in vitro, allowing recordings to be obtained from brain slices at sites deeper within the tissue than currently available surface recording methods permit. However, existing recording chambers do not allow for the insertion of lamina-spanning probes that enter through the edges of brain slices. Here, we present a novel brain slice recording chamber design that accomplishes this goal. The device provides a stable microfluidic perfusion environment in which tissue health is optimized by superfusing both surfaces of the slice. Multichannel electrodes can be inserted parallel to the surface of the slice, at any depth relative to the surface. Access is also provided from above for the insertion of additional recording or stimulating electrodes. We illustrate the utility of this recording configuration by measuring current sources and sinks during theta burst stimuli that lead to the induction of long-term potentiation in hippocampal slices., ((c) 2010 Elsevier B.V. All rights reserved.)