Intracellular production of a nuclease
View Patent ↗Methods and compositions are provided that relate to obtaining a recombinant DNA and RNA cleaving nuclease. This involves the over-expression of a fusion protein between maltose-binding protein and a truncated nuclease in a soluble form in the cytoplasm of a host cell from which it can be readily extracted.
1. A DNA segment encoding a recombinant fusion protein containing maltose-binding protein fused to a nuclease comprising SEQ ID NO: 2 or SEQ ID NO: 4.
2. A host cell comprising a DNA segment encoding a recombinant fusion protein containing maltose-binding protein fused to a nuclease comprising SEQ ID NO: 2 or SEQ ID NO: 4.
3. A method for obtaining a nuclease, comprising:
(a) obtaining a DNA segment encoding a recombinant fusion protein containing maltose-binding protein fused to a nuclease comprising SEQ ID NO: 2 or SEQ ID NO: 4;
(b) expressing the DNA segment within a host cell to produce the fusion protein in the cytoplasm;
(c) lysing the host cell; and
(d) obtaining the isolated nuclease.
4. The method of claim 3 , wherein the nuclease comprises SEQ ID NO: 2.
5. The method of claim 3 , wherein the nuclease comprises SEQ ID NO: 4.
6. The DNA segment of claim 1 , wherein the nuclease comprises SEQ ID NO: 2.
7. The DNA segment of claim 1 , wherein the nuclease comprises SEQ ID NO: 4.
8. The host cell of claim 2 , wherein the nuclease comprises SEQ ID NO: 2.
9. The host cell of claim 2 , wherein the nuclease comprises SEQ ID NO: 4.