Team:CSMU Taiwan/Part
Part
Overview
After we complete designing the whole experiments, we start getting into designing the sequences that are used for synthesis of the circular probe or our kit GotCha. Our dry lab uses the python language to build a model which can formulate sequences of the circular primer, and then we can check the thermal dynamic stability and secondary structures on the online secondary structure prediction websites to decide which DNA sequence we should choose (For the python program, please check our Model).
The following shows the parts that make up our Circular DNA and Immobilisation Probe：
Immobolization probe
This DNA chain composes an Anchor Sequence and Circular DNA Binding site, responsible for the securing the Circular DNA probe to magnetic beads.
Circular DNA Binding site
It is composed of 7 nucleotides complementary to Circular DNA, and being used to form the bond between the Immobolization probe and the Circular DNA.
Anchor Sequence
A 28nt long dna sequence, containing a poly-A-tail like segment, and modified with biotin modification at poly A tail end in order to adhere to magnetic beads.
Circular DNA
This part composes an Immobolization Probe Binding Site, MicroRNA Binding Site and Functionless Sequence, with a total length of 75 nt. It is the Circular DNA Probe used for Rolling Circle Amplification.
Immobolization Probe Binding Site
Connect to Immobolization probe, which has 7 nucleotides complementary to Immobolization probe.
MicroRNA Binding Site
It is the location where the target microRNA is captured through complementary base pairing to the target microRNA. It will mark the starting point for the phi29 polymerase to start the RCA reaction as target microRNAs are treated as the primer.
Functionless Sequence
It is divided into two blocks, 6 nucleotides are located between Immobolization Probe Binding Site and MicroRNA Binding Site, and the other section of Functionless Sequence is 39 nucleotides long and connects the rest section of Circular DNA.
Single Part
Name |
Description |
Length |
|---|---|---|
Circular DNA Binding site |
7nt |
|
Immobolization Probe Binding Site |
7nt |
|
miR-664a-3p binding site |
23nt |
|
miR-33b-3p binding site |
22nt |
|
Anchor Scquence |
28nt |
Composite Part
Name |
Description |
Length |
|---|---|---|
Immobolization Probe |
35nt |
|
Circular DNA corresponding to target miR-664a-3p |
75nt |
|
Circular DNA corresponding to target miR-664a-3p |
75nt |
|
Circular DNA corresponding to target miR-664a-3p |
75nt |
|
Circular DNA corresponding to target miR-664a-3p |
75nt |
|
Circular DNA corresponding to target miR-664a-3p |
75nt |
|
Circular DNA corresponding to target miR-664a-3p |
75nt |
|
Circular DNA corresponding to target miR-33b-3p |
75nt |
|
Circular DNA corresponding to target miR-33b-3p |
75nt |
|
Circular DNA corresponding to target miR-33b-3p |
75nt |
|
Circular DNA corresponding to target miR-33b-3p |
75nt |
|
Circular DNA corresponding to target miR-33b-3p |
75nt |
|
Circular DNA corresponding to target miR-33b-3p |
75nt |
