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Vivo-Morpholinos are also effective and easy to use in cultures.
This property can be exploited to block translation, block splicing, block miRNAs or their targets, and block ribozyme activity.
By sterically blocking the translation initiation complex, Morpholinos can knock down expression of many target sequences completely enough that after waiting for existing protein to degrade, the target protein band disappears from Western blots.
Morpholinos delivered to rat neonatal cardiomyocytes in cell culture using Endo-PorterKrautbauer S, Eisinger K, Neumeier M, Hader Y, Buettner R, Schmid PM, Aslanidis C, Buechler C.
All Vivo-Morpholinos are sterilized.
[Epub ahead of print]
miRNA target-protecting Morpholino delivered to cultured cardiomyocytes with Endo-PorterWang P, Calise J, Powell K, Divald A, Powell SR.
If synthesis of NFs is important for axon regeneration in the CNS, then inhibiting NF synthesis should inhibit regeneration. In the present study we therefore examined, the effect of NFs on axon regeneration by blocking the translation of a key NF subunit, NF180, with antisense morpholino oligonucleotides (MOs). We also evaluated: 1) Whether, in addition to regrowth from severed axons (true regeneration), there is collateral sprouting from spare axons after hemisection of lamprey spinal cord; 2) NFs protein expression and time course of expression in lampreys of different sizes (and therefore different ages) for up to 1 year after spinal cord TX. These features of the response to spinal cord injury are very important for using lamprey as a model for axonal regeneration and for using expression of NFs as a marker of axonal regeneration.
Vivo-Morpholinos are very simple to use in cultured cells.
Vivo‐Morpholino injections in adult zebrafish: "Briefly, zebrafish were anesthetized in tricaine 0.05 mg/mL, and 2 μL of 0.1 mmol/L vivo‐Mo solution was loaded in a glass capillary and injected with a standard microinjector (IM300 Microinjector; Narishige, Tokyo, Japan) on days 0, 2, 4, 6, 8, 10, 12, and 14."
Paveliev M, Fenrich KK, Kislin M, Kuja-Panula J, Kulesskiy E, Varjosalo M, Kajander T, Mugantseva E, Ahonen-Bishopp A, Khiroug L, Kulesskaya N, Rougon G, Rauvala H.
2017;[Epub ahead of print] doi:10.1523/JNEUROSCI.3717-16.2017
adult male Sprague Dawley rats injected with Vivo-Morpholino in PBS into nucleus accumbens, 30 pmol/side in 1µl though 33 gauge microinjectors fit though cannulae at 0.5µl/min, daily for three days.
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All morpholinos were at a concentration of 500 nM per 1.0 μl.
The sea lamprey has been used as a model for the study of axonal regeneration after spinal cord injury. Previous studies have suggested that, unlike developing axons in mammal, the tips of regenerating axons in lamprey spinal cord are simple in shape, packed with neurofilaments (NFs), and contain very little F-actin. Thus it has been proposed that regeneration of axons in the central nervous system of mature vertebrates is not based on the canonical actin-dependent pulling mechanism of growth cones, but involves an internal protrusive force, perhaps generated by the transport or assembly of NFs in the distal axon. In order to assess this hypothesis, expression of NFs was manipulated by antisense morpholino oligonucleotides (MO). A standard, company-supplied MO was used as control. Axon retraction and regeneration were assessed at 2, 4 and 9 weeks after MOs were applied to a spinal cord transection (TX) site. Antisense MO inhibited NF180 expression compared to control MO. The effect of inhibiting NF expression on axon retraction and regeneration was studied by measuring the distance of axon tips from the TX site at 2 and 4 weeks post-TX, and counting the number of reticulospinal neurons (RNs) retrogradely labeled by fluorescently-tagged dextran injected caudal to the injury at 9 weeks post-TX. There was no statistically significant effect of MO on axon retraction at 2 weeks post-TX. However, at both 4 and 9 weeks post-TX, inhibition of NF expression inhibited axon regeneration.
with nontargeting control vivo-morpholino ...
with 12.5 mg/Kg vivo-morpholino for two consecutive days at an interval of 24 hr
de Morrée A, van Velthoven CTJ, Gan Q, Salvi JS, Klein JDD, Akimenko I, Quarta M, Biressi S, Rando TA.
or c-Src-targeting vivo-morpholino ...
A functional role for this gene expression in the angiogenic switch was then shown using antisense morpholino oligonucleotides to suppress the increased COL1A2 expression in TSP1 null explants.
[Epub ahead of print]Morpholino delivery into fibroblast cultures.
Epub 2017 Aug 28.
Vivo-Morpholinos combined with Endo-Porter in VCaP (ATCC CRL-2876) cell culture
Lu Y, Hu Z, Mangala LS, Stine ZE, Hu X, Jiang D, Xiang Y, Zhang Y, Pradeep S, Rodriguez-Aguayo C, Lopez-Berestein G, Demarzo A, Sood AK, Zhang L, Dang CV.
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