i-Fect Delivers Plasmids!

Important for Gene Expression Studies.
I have posted many examples of how our customers use i-Fect^TM  and other Transfection Solutions for Gene Manipulation Studies. There are also many publications.

Here we feature how i-Fect was used to delivery plasmids to the CNS: Sara Elramah, María José López-González, Matthieu Bastide, Florence Dixmérias, Olivier Roca-Lapirot, Anne-Cécile Wielanek-Bachelet, Anne Vital, Thierry Leste-Lasserre, Alexandre Brochard, Marc Landry & Alexandre Favereaux. Spinal miRNA-124 regulates synaptopodin and nociception in an animal model of bone cancer pain. Scientific Reports 7, Article number: 10949 (2017) doi:10.1038/s41598-017-10224-1...Intrathecal administration of miRNAs and ShRNA To over-express miR-124, we cloned the pre-miRNA sequence of miR-124 into a plasmid. To determine cells expressing this miR-124 encoding plasmid, we added a GFP-coding sequence to the construct under the control of an IRES. Thus, miR-124 over-expressing cells also express GFP. To inhibit synaptopodin expression, we cloned a ShRNA sequence directed against synaptopodin into a plasmid. To determine cells expressing this ShRNA, we added a GFP-coding sequence to the construct under the control of an IRES. Thus, ShRNA expressing cells also expressed GFP. Two micrograms of these plasmids or the corresponding controls, were solubilized in 10 µl of i-Fect reagent (Neuromics, Edina, USA), and injected intrathecally between the L5 and L6 lumbar vertebrae every two days for a total of 3 injections, according to the manufacturer’s instructions and previously published experiments...

Figures: (C and D) Immunostaining of synpo in spinal cord after miR-124 intrathecal injections: only the dorsal horn which receive nociceptive information was quantified (white dash area). Measurement of synaptopodin stained area reveals ability of miR-124 to inhibit endogenous Synpo expression (20/3 and 17/3 denotes number of sections/animals for control and miR-124-injected mice, respectively.

I am confident there will be many more positive reports regarding our Transfection Reagents.

Protein Purification Solutions

Strep-tag®Tools + Strep-Tactin Columns

We are seeing a notable uptick in use of our Protein Purification Tools. This includes vectors and purification columns. We are especially delighted with the increased use of of our Strep-Tag + Strep-Tactin Columns.

Figure: Strep-Tag+Strep-Tactin Methods

Here's an example of researchers using the technology to purify heme proteins for the study of the diptheria pathogen: Courtni E. Allen and Michael P. Schmitt. Utilization of Host Iron Sources by Corynebacterium diphtheriae: Multiple Hemoglobin-Binding Proteins Are Essential for the Use of Iron from the Hemoglobin-Haptoglobin Complex. J. Bacteriol. February 2015 vol. 197 no. 3 553-562. doi: 10.1128/JB.02413-14...Cell debris was removed by centrifugation at 12,000 × g at 4°C, and the supernatant fraction containing the soluble protein was purified on Strep-Tactin columns (Neuromics) for Strep-tagged constructs...

Figure: (A) HtaA and derivatives were assessed for the ability to bind Hb-Hp by ELISA. GST was included as a negative control. Experiments were repeated at least three times, with similar results. Results of a representative experiment are shown. (B) Binding of HtaA and various derivatives to Hb-Hp at a 200 nM concentration is shown. Binding of HtaA-Y361A and CR2-Y361A to Hb-Hp was significantly different from binding with HtaA-wt and CR2, respectively (**, P less than 0.0001). Binding of CR1 to Hb-Hp was significantly different from that with HtaA-wt (***, P less than 0.05). *, HtaA does not bind Hp (HtaA-Hp). Values represent the means from three independent experiments (±SD).

Looking for protein purification solutions? Do not hesitate to contact me: 612-801-1007 or pshuster@neuromics.com.