Hydrodynamic Tail Injection Process
1. Materials
Materials |
Size |
Details |
Manufacturer |
Trade marker |
---|---|---|---|---|
Millex filter |
0.22um |
Slmp025ss, quantity 50pk |
Millipore |
Millex |
Needle |
25Gx5/8 |
100, REF 305122 |
BD |
BD precisionGlide™ |
Needle |
26Gx5/8 Sub-Q |
100, REF 305115 |
BD |
BD precisionGlide™ |
Syringe |
3ml |
REF 309657 |
BD |
Luer-lok™ tip |
Syringe |
10ml |
REF 309604 |
BD |
Luer-lok™ tip |
Saline |
||||
tube |
50ml |
BD |
||
MICE fix container |
TV-150 |
Braintree scientific INC ,MA 02185 |
||
Red-ray Lamp |
2. Tail injection
2.1 plasmid solution preparation
- In most cases, we inject 20ug of “oncogene” into the mice, and SB is 1/25 of the total oncogene injected. For example, for AKT/Ras model, we need 20ug AKT, 20ug Ras and 1.6ug SB for each mouse.
- Calculate the plasmid needed to use in according to the dosage for the experimental design. Usually used 2ml/mouse (weight is about 20g when mice are at 6 weeks old), and the total volume is equal to (n+1) x2 ml, such as the mice number is 4, then need to prepare solution 10ml, namely (4+1)x2=10
- Filter the plasmids, after diluted the plasmid to the final volume, and mix well by shaking upside, then filter the solution to a new 50ml tube by filtering with 0.22ul filter. Be careful not contaminated with other plasmids when more than 2 groups experiments. Marked the name and plasmids information on the tube.
2.2 tail injection
- Take out 3ml syringe, load 2ml plasmid solution.
- Take one mouse from the cage, open the red-ray heater, and put the mouse tail under the heater for about 10-20 seconds, make the tail vein can easily find.
- Fix the mice to the mice container, make mouth up, so that the mice do not easily move in the fixer, give the mice the plasmid through tail vein within 5-8 seconds.
Note
1. about plasmid preparation
- When the plasmid concentration is too low, usually below 0.1ug/ml, then need to concentration, or deduced the volume from the total volume for tail injection.
- Fresh prepared the plasmid solution for tail injection in the same day is better.
- Be avoiding of frozen- thawing plasmids too many times.
For Maxi prep, we found the following kit is the best:
Plasmid Maxiprep Kit :GenElute™ HP Endotoxin-Free Plasmid Maxiprep Kit; Part Number: NA0410
2. about injection
- Usually keep the slope of the needle up when tail injection.
- Make sure there is no bubble in the syringe.
- If keep the needle slope in parallel with the outside part of the syringe, it is convenient for the tail injection.