Method for producing a transgenic pig using a hyper-methylated transposon

• US 8,309,791 B2
• Filed: 07/16/2009
• Issued: 11/13/2012
• Est. Priority Date: 07/16/2008
• Status: Expired due to Fees

• Current Assignee: Recombinetics, Inc
• Original Assignee: Recombinetics, Inc
• Inventors: Fahrenkrug, Scott C., Carlson, Daniel F., Geurts, Aron M.
• Primary Examiner(s): Crouch, Deborah
• Application Number:
  US12/504,364
• Publication Number:
  US 20100146655A1
• Time in Patent Office: 1,216 Days
• Field of Search: None
• US Class Current: 800/17
• CPC Class Codes: A01K 2217/058 due to expression of inhibitory nucleic acid, e.g. siRNA, antisense
• CPC Class Codes: A01K 2217/203 Animal model comprising inducible/conditional expression system, e.g. hormones, tet
• CPC Class Codes: A01K 2227/101 Bovine
• CPC Class Codes: A01K 2227/103 Ovine
• CPC Class Codes: A01K 2227/105 Murine
• CPC Class Codes: A01K 2227/108 Swine
• CPC Class Codes: A01K 2267/03 Animal model, e.g. for test or diseases
• CPC Class Codes: A01K 2267/035 Animal model for multifactorial diseases
• CPC Class Codes: A01K 67/0273 Cloned vertebrates
• CPC Class Codes: A01K 67/0275 Genetically modified vertebrates, e.g. transgenic
• CPC Class Codes: A61D 19/00 Instruments or methods for reproduction or fertilisation
• CPC Class Codes: C07K 14/4712 Cystic fibrosis
• CPC Class Codes: C12N 15/1138 against receptors or cell surface proteins
• CPC Class Codes: C12N 15/8509 for producing genetically modified animals, e.g. transgenic
• CPC Class Codes: C12N 2015/8527 for producing animal models, e.g. for tests or diseases
• CPC Class Codes: C12N 2310/14 interfering N.A.
• CPC Class Codes: C12N 2800/30 Vector systems comprising sequences for excision in presence of a recombinase, e.g. loxP or FRT
• CPC Class Codes: C12N 2800/90 Vectors containing a transposable element

Abstract

Transgenic artiodactyls are described as well as methods of making and using such artiodactyls.

Claims

1. A method for making a transgenic pig, said method comprising:

preparing a pig egg with an in vitro hypermethylated transposon that comprises a transcriptional unit comprising a regulatory region operably linked to a target nucleic acid sequence, said transposon being inserted into a genome of the egg;

transferring said egg to a recipient female; and

allowing said egg to develop in said recipient female to produce said transgenic pig, wherein the pig is gestated in the female and expresses the target nucleic acid sequence.

2. The method of claim 1 wherein preparing the pig egg comprises introduction of the transposon and a transposase into a single-cell fertilized egg, with the transposase inserting the transposon into the genome.

3. The method of claim 1 wherein preparing the pig egg comprises enucleation of the egg followed by transfer of a nucleus of a somatic pig cell into the egg, with the transferred nucleus comprising the gene and the transposon inserted into the genome.

4. The method of claim 1 wherein preparing the pig egg comprises transfer of chromatin of a somatic pig cell into the egg, with the chromatin of the somatic cell comprising the transposon.

5. The method of claim 1 wherein the hypermethylated transposon comprises hypermethylation between inverted terminal repeats of the transposon.

6. The method of claim 1 wherein all cytosine-phosphodiester-guanine sites in the transposon are methylated.

7. The method of claim 1 wherein at least 90% of cytosine-phosphodiester-guanine sites in the transposon are methylated.

8. The method of claim 1 wherein said transposon is selected from the group consisting of Sleeping Beauty, Tol2, PiggyBac, Frog Prince, Minos, and Hsmar1.