Rolling circle amplification (RCA) assay needs a circular form of single-stranded DNA and a primer that will be turn on by a phi29 DNA polymerase. A syringe is able to connect to one end of the adapter and a P200 tip can fit onto the other end. We’ve developed a Luer-lock adapter by 3D printing, in which a mini Nickel column was embedded that was used to capture His-tagged phi29 DNA polymerase. The His-phi29 DNA polymerase will be produced by the phage-infected Salmonella. Salmonella phages were genetically engineered to equip with phi29 DNA polymerase gene having a N-terminal His tag. The limit of detection with our kit was described in detail on our page of PROOF-OF-CONCEPT.
If Salmonella exists, the signal will be observed or vice versa. Read data by measuring the signal at Ex/Em=500/530 nm. Slightly shake the plate to mix the DNA and dye. Push the plunger of the syringe to elute the single-stranded DNA (ssDNA) as RCA products into a well of a 96-well black microplate containing a lyophilized fluorescent DNA-binding dye such as EvaGreen® Dye.ħ. Incubate for 30 min at room temperature.Ħ. Perform RCA in the Ni column that captures His-Phi29 DNA polymerase if any. The reagent consisting of a circular DNA and a primer in Vial B is drew onto the column in the Luer adapter using a P200 tip.
#SNAPGENE ADAPTORS FULL#
Let the mixtures pass through the column to capture any His-tagged proteins (Optional) If the sample is full of proteins or complicated contents, try to wash the column 1-3 times with the Wash Buffer in Via C we provided.ĥ. The Luer adapter with Ni-NTA column is locked onto the syringe. Mix and incubate for 25 minutes at room temperature.Ĥ. The engineered phage liquid in Vial A is drawn into the syringe containing the sample. Put liquid samples (if your sample is solid, first make it liquid by mixing with some water) into any kind of syringe or draw the samples with the syringe.ģ. Samples to be examined including eggs and egg-related products are collected in a container (e.g., a beaker of any size, better smaller than 50ml)Ģ.
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