AssemblX: a user-friendly toolkit for rapid and reliable multi-gene assemblies

The assembly of large DNA constructs coding for entire pathways poses a major challenge in the field of synthetic biology. Here, we present AssemblX, a novel, user-friendly and highly efficient multi-gene assembly strategy. The software-assisted AssemblX process allows even unexperienced users to rapidly design, build and test DNA constructs with currently up to 25 functional units, from 75 or more subunits. At the gene level, AssemblX uses scar-free, overlap-based and sequence-independent methods, allowing the unrestricted design of transcriptional units without laborious parts domestication. The assembly into multi-gene modules is enabled via a standardized, highly efficient, polymerase chain reaction-free and virtually sequence-independent scheme, which relies on rare cutting restriction enzymes and optimized adapter sequences. Selection and marker switching strategies render the whole process reliable, rapid and very effective. The assembly product can be easily transferred to any desired expression host, making AssemblX useful for researchers from various fields

Feasibility studies of multimodal nonlinear endoscopy using multicore fiber bundles for remote scanning from tissue sections to bulk organs

Abstract Here, we report on the development and application of a compact multi-core fiber optical probe for multimodal non-linear imaging, combining the label-free modalities of Coherent Anti-Stokes Raman Scattering, Second Harmonic Generation, and Two-Photon Excited Fluorescence. Probes of this multi-core fiber design avoid moving and voltage-carrying parts at the distal end, thus providing promising improved compatibility with clinical requirements over competing implementations. The performance characteristics of the probe are established using thin cryo-sections and artificial targets before the applicability to clinically relevant samples is evaluated using ex vivo bulk human and porcine intestine tissues. After image reconstruction to counteract the data’s inherently pixelated nature, the recorded images show high image quality and morpho-chemical conformity on the tissue level compared to multimodal non-linear images obtained with a laser-scanning microscope using a standard microscope objective. Furthermore, a simple yet effective reconstruction procedure is presented and demonstrated to yield satisfactory results. Finally, a clear pathway for further developments to facilitate a translation of the multimodal fiber probe into real-world clinical evaluation and application is outlined

The relation between soluble apoptotic proteins and subclinical cardiotoxicity in adjuvant-treated breast cancer patients

Background: Circulating apoptotic proteins are increased in heart failure patients. We evaluated whether circulating soluble apoptosis-related protein levels change after anthracycline-containing chemotherapy and radiotherapy in relation to cardiac dysfunction or the applied treatment. Patients and Methods: Circulating apoptotic proteins were measured with immunoassay in 40 breast cancer patients following surgery (T0), one month (T1) and one year (T2) after epirubicin-based chemotherapy. Standard-dose (n=21) or high-dose (n=19) myeloablative chemotherapy, preceded irradiation and tamoxifen. Circulating apoptotic proteins were compared with previous cardiac evaluations. Results: Soluble tumor necrosis factor receptor 1 (+30%), 2 (+43%) and Fas (+40%) were transiently increased at T-1 compared to T-0, whereas Fas ligand (-64%) was transiently decreased, especially in the high-dose group. Apoptosis markers were not associated with cardiac dysfunction. Conclusion: Significant, but transient changes in soluble apoptotic protein levels were observed, particularly after high-dose chemotherapy. No relation was found between apoptosis-related proteins and cardiotoxicity