Methods and materials for assembling nucleic acid constructs
AssigneeMayo Foundation for Medical Education and Research
InventorsStephen C. Ekker, Karl J. Clark, Jarryd M. Campbell, Chun Hang Alvin Ma
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Abstract
This document provides methods and materials for assembling nucleic acid constructs (e.g., TALENs). For example, methods for assembling TALEs that are rapid, flexible for use in many cloning scaffolds (such as common nuclease and nickase backbones), and achievable with standard molecular biology laboratory tools, thereby making TALEs a more accessible genome system, are provided.
Claims17
A method for assembling a polynucleotide sequence that encodes a transcription activator-like effector (TALE)-DNA modifying enzyme that comprises
- (A) a TALE binding domain comprising a plurality of repeat modules, and
- (B) a DNA modifying enzyme domain,
- wherein the method comprises combining, in a single reaction mixture,
- (a) a plurality of DNA segments, wherein an individual DNA segment of said plurality of DNA segments encodes a repeat module of said TALE binding domain repeat modules, and wherein said plurality of DNA segments comprises at least two trimer DNA sequences, and
- (c) a first expression vector encoding said DNA modifying enzyme domain
- wherein said combining is under conditions so as to generate a second expression vector encoding said polynucleotide sequence encoding said TALE-DNA modifying enzyme in a single cloning reaction.
A kit comprising a first plurality of wells that contain 64 trimer repeat modules of a TALE binding domain, wherein each of said plurality of wells contains one trimer repeat module of said 64 trimer repeat modules.
Dependencies are inferred from the claim text.