By Grigory S Filonov, Samie Jaffrey
Methods in Enzymology: Visualizing RNA Dynamics within the Cell maintains the legacy of this premiere serial with caliber chapters authored via leaders within the box. This quantity covers study equipment visualizing RNA dynamics within the phone, and contains sections on such subject matters as identity of RNA cis-regulatory sequences, IRAS, IMAGEtags, MERFISH, plant RNA labeling utilizing MS2, and visualization of 5S dynamics in stay cells utilizing photostable corn probe.
- Continues the legacy of this preferable serial with caliber chapters authored by way of leaders within the box
- Covers study tools in visualizing RNA dynamics within the cell
- Contains sections on such issues as id of RNA cis-regulatory sequences, IRAS, IMAGEtags, MERFISH, plant RNA labeling utilizing MS2 and visualization of 5S dynamics in reside cells utilizing photostable corn probe
Read Online or Download Visualizing RNA Dynamics in the Cell PDF
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Extra resources for Visualizing RNA Dynamics in the Cell
This algorithm will create an N-bit binary barcode for each of these spots. Step 2: Construct a barcode for each found spot. For each spot in the list created in Step 1, compute the distance to the nearest spot in each round of hybridization. This will create a vector of N distances. To convert this vector into a binary barcode, compare the ith distance to the maximum allowed distance (a parameter of this algorithm). If this distance is less than or equal to this maximum distance, assign a “1” to the ith bit of the barcode for this spot.
Place this box into a 37°C incubation chamber and incubate for at least 12 h. In some cases, we find that longer incubations ($36 h) increase the quality of staining. Again, we recommend varying the incubation time for each probe set to identify the optimal incubation time. Step 4: Wash away residual encoding probes. Preheat 6 mL of encoding probe wash buffer per sample to 47°C in a water bath. Slowly peel the coverslip off of the microscope slide taking care not to crack the coverslip. If it appears to be stuck, immerse the assembly in a layer of encoding probe wash buffer for 5 min to loosen.
For example, imagine that one “blank” word is counted more frequently than a barcode representing a low-abundance RNA. If these two barcodes are separated by a large HD, ie, these barcodes have very few “1” bits in common, then it is possible that the errors that produced observations of this “blank” barcode may not be relevant for the measurement of the barcode associated with the low-abundance RNA. To partially capture the relationship between different barcodes and improve estimates of the accuracy of different barcode measurements, we have developed a metric that we term the confidence ratio.