Micro-droplets for cell-free protein synthesis and preparation method thereof
A protein synthesis and microdroplet technology, which is applied to the field of microdroplets for cell-free protein synthesis and its preparation, can solve the problems of complicated operation, difficult to realize the simple and efficient synthesis of cell-free protein, difficult separation and purification, etc. Simple and fast method
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Embodiment 7
[0047] Preparation of cell disruption products:
[0048] The cultured Escherichia coli (Escherichia coli BL21(DE3) CICC 23796, purchased from CICC) was collected, suspended, centrifuged, mechanically broken, and stored.
[0049] In addition to mechanical methods, cell disruption methods can also be repeated freezing and thawing methods, ultrasonic treatment methods, enzymatic methods, alkaline lysis methods or chemical permeation methods.
[0050] Eukaryotic cells can also be used for disruption.
[0051] Examples 8-10 are composition 1, see Table 3.
[0052] table 3
[0053]
Example 8
Example 9
Example 10
Cell disruption products
Example 7 Preparation of 1g
Example 7 Preparation of 1g
Example 7 Preparation of 1g
Example 1 Preparation of 0.1ml
Example 2 Preparation of 0.05ml
Example 3 Preparation of 0.01ml
Example 4 Preparation of 3ml
Example 5 Preparation of 2ml
Example 6 Preparation of 1ml
[0054] Examples 11-13 are amino acid m...
Embodiment 20-22
[0063] Examples 20-22 are composition 2 (each component is volume ratio), see Table 7
[0064] Table 7
[0065]
Example 20
Example 21
Example 22
Amino acid mixture
8mL prepared in Example 11
5mL prepared in Example 12
2mL prepared in Example 13
Reaction buffer
30mL prepared in Example 14
25mL Example 15 preparation
20mL prepared in Example 16
Energy supplement
5mL prepared in Example 17
3mL prepared in Example 18
2mL prepared in Example 19
Ribonucleic acid polymerase aqueous solution
1mL 150μg / ml
1mL 70μg / ml
1mL 5μg / ml
[0066] Gene plasmids can be purchased or obtained through genetic modification.
[0067] pRset-CFP (purchased from: Promega Corporation)
[0068] pRset-YFP (purchased from: Promega Corporation)
[0069] pRset-eGFP (purchased from: Promega Corporation)
[0070] The above examples of plasmids are intended to enable those skilled in the art to better understand the present invention, but they are not limited, and other plasmids can also be used in ...
Embodiment 23-25
[0071] Examples 23-25 are oil phase
[0072] Table 8
[0073]
[0074] EM 90 (Degussa)
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