Microfluidic platforms use controlled fluid flows to provide physiologically relevant biochemical and biophysical cues to cultured cells in a well-defined and reproducible manner. Undisturbed flows are critical in these systems, and air bubbles entering microfluidic channels can lead to device delamination or cell damage. To prevent bubble entry into microfluidic channels, we report a low-cost, Rapidly Integrated Debubbler (RID) module that is simple to fabricate, inexpensive, and easily combined with existing experimental systems. We demonstrate successful removal of air bubbles spanning three orders of magnitude with a maximum removal rate (dV/dt)max = 1.5 mL min−1, at flow rates required to apply physiological wall shear stress (1–200 dyne cm−2) to mammalian cells cultured in microfluidic channels.

Cyanobacteria produce structurally and functionally diverse polyketides, nonribosomal peptides and their hybrids. Sfp-type phosphopantetheinyl transferases (PPTases) are essential to the production of these compounds via functionalizing carrier proteins (CPs) of biosynthetic megaenzymes. However, cyanobacterial Sfp-type PPTases remain poorly characterized, posing a significant barrier to the exploitation of cyanobacteria for biotechnological and biomedical applications. Herein, we describe the detailed characterization of multiple cyanobacterial Sfp-type PPTases that were rationally selected. Biochemical characterization of these enzymes along with the prototypic enzyme Sfp from Bacillus subtilis demonstrated their varying specificities toward 11 recombinant CPs of different types of biosynthetic pathways from cyanobacterial and Streptomyces strains. Kinetic analysis further indicated that PPTases possess the higher binding affinity and catalytic efficiency toward their cognate CPs in comparison with noncognate substrates. Moreover, when chromosomally replacing the native PPTase gene of Synechocystis sp. PCC6803, two selected cyanobacterial PPTases and Sfp supported the growth of resulted mutants. Cell lysates of the cyanobacterial mutants further functionalized recombinant CP substrates. Collectively, these studies reveal the versatile catalysis of selected cyanobacterial PPTases and provide new tools to synthesize cyanobacterial natural products using in vitro and in vivo synthetic biology approaches.

Microfluidic platforms use controlled fluid flow to provide physiologically relevant biochemical and biophysical cues to cultured cells in a well-defined and reproducible manner. In these systems, undisturbed flows are critical and air bubbles entering microfluidic channels can result in device delamination or cell damage. To prevent bubble entry, we report a low-cost, Rapidly Integrated Debubbler (RID) module that is simple to fabricate, inexpensive, and easily combined with existing experimental systems. We demonstrate successful removal of air bubbles spanning three orders of magnitude with a maximum removal rate (dV/dt)max = 1.5 mL min -1 , at flow rates corresponding to physiological fluid-induced wall shear stresses (WSS) needed for biophysical stimulation studies on cultured mammalian cell populations.