PCR is now a common and often indispensable technique used in medical laboratory and clinical laboratory research for a broad variety of applications including biomedical research and criminal forensics.
Provided are vectors and vector systems, some of which encode one or more components of a CRISPR complex, as well as methods for the design and use of such vectors.
More specifically, all referenced documents are incorporated by reference to the same extent as if each individual document was specifically and individually indicated to be incorporated by refere ce. The government has certain rights in the invention. Functional genomics is a field of molecular biology that may be considered to utilize the vast wealth of data produced by genomic projects such as genome sequencing projects to describe gene and protein functions and interactions.
Contrary to classical genomics, functional genomics focuses on the dynamic aspects such as gene transcription, translation, and protein-protein mteractions, as opposed to the static aspects of the genomic information such as DNA sequence or structures, though these static aspects are very important and supplement one's understanding of cellular and molecular mechanisms.
Functional genomics attempts to answer questions about the function of DN A at the levels of genes, RNA transcripts, and protein products.
A key characteristic of functional genomics studies is a genome-wide approach to these questions, generally involving ig -throughput methods rather than a more traditional "gene-by-gene" approach. Given the vast inventory of genes and genetic information it is advantageous to use genetic screens to provide information of what these genes do, what cellular pathways they are involved in and how any alteration in gene expression can result in a particular biological process.
There are three key elements for a functional genomics screen: Good reagents that allow for precise genome targeting technologies are needed to enable systematic reverse engineering of causal genetic variations by allowing selective perturbation of individual genetic elements, as well as to advance synthetic biology, biotechnological, and medical applications.
Adding the CRISPR-Cas system to the repertoire of genome sequencing techniques and analysis methods may significantly simplify the methodology and accelerate the ability to catalog and map genetic factors associated with a diverse range of biological functions and diseases.
Aspects of this invention address this need and provide related advantages. The guide sequence is linked to a tracr mate sequence, which in turn hybridizes to a tracr sequence. This library may potentially comprise guide RNAs that target each and eve ' gene in the genome of an organism.
In some embodiments of the invention the organism or subject is a eukaryote including mammal including human or a non-human eukaryote or a non-human animal or a non-human mammal. In some embodiments, the organism or subject is a non-human animal, and may be an arthropod, for example, an insect, or may be a nematode.
In some methods of the invention the organism or subject is a plant. In some methods of the invention the organism or subject is a mammal or a non-human mammal.
A non-human mammal may be for example a rodent preferably a mouse or a ratan ungulate, or a primate. In some methods of the invention the organism or subject is algae, including microa! In an embodiment of the invention, the Cas protein is a Cas9 protein.
In another embodiment, the one or more vectors are plasmid vectors. In a furher embodiment, the regulatory element operably linked to the Cas protein is an inducible promoter, e.
The invention comprehends that the population of cells is a population of eukaryotic cells, and in a preferred embodiment, the population of cells is a population of embryonic stem ES cel ls.Inventor Feng Zhang Neville Espi SANJANA Ophir SHALEM Original Assignee The Broad Institute, Inc.
Massachusetts Institute Of Technology Priority date (The priority . Search the history of over billion web pages on the Internet. Co-incubation progressed for two or plasmids using EcoRI restriction enzyme, and ligated using T4 DNA three days at room temperature, with the plates maintained with- ligase (New England Biolabs, Ipswich, MA) into the EcoRI site of out Parafilm.
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Msc. Biotechnology Industrial biotechnology or microbial biotechnology is the ing on the enzyme involved: More than restriction endonucleases are known in bacteria.5/5(1).
Study revealed potential roles for some open reading frames of previously unknown function. Akerley BJ, et al. A genome-scale analysis for identification of genes required for growth or survival of haemophilus influenzae.
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