Abstract: The disclosure relates generally to genetic manipulation of stem and primary cells and to reprogramming of somatic cells, more specifically, human cells. In particular, compositions and methods are disclosed for the generation and maintenance of such engineered cells.

Abstract: The disclosure relates generally to stem cell biology and more specifically to genetic manipulation of stem cells. Methods and compositions using recombinational cloning techniques are disclosed which allow the construction and insertion of complex genetic constructs into embryonic and adult stem cells and progenitor cells. The methods disclosed will allow the harvesting of adult stem cells pre-engineered with integration sites to facilitate early passage genetic modification.

Abstract: The present invention is in the fields of biotechnology and molecular biology. More particularly, the present invention relates to cloning or subcloning one or more nucleic acid molecules comprising one or more type IIs restriction enzyme recognition sites. The present invention also embodies cloning such nucleic acid molecules using recombinational cloning methods such as those employing recombination sites and recombination proteins. The present invention also relates to nucleic acid molecules (including RNA and iRNA), as well as proteins, expressed from host cells produced using the methods of the present invention.

Abstract: Embodiments are directed to devices and methods for processing, cultivating or otherwise manipulating cell cultures which may be disposed on a flat or substantially flat surface such as cell culture substrate material. Devices and methods are disclosed for dividing a cell culture layer into divided portions, including isolated divided portions, that may then be transferred from the cell culture to a new location. For some embodiments, the divided portions may be transferred to a new cell culture support substrate in order to continue to grow and cultivate the cell line.

Abstract: A method of generating a double stranded (ds) recombinant nucleic acid molecule covalently linked in both strands by contacting two or more ds nucleotide sequences with a topoisomerase under conditions such that both termini of at least one end of a first ds nucleotide sequence are covalently linked by the topoisomerase to both termini of at least one end of a second ds nucleotide sequence is provided. Also provided is a method for generating a ds recombinant nucleic acid molecule covalently linked in one strand, by contacting two or more ds nucleotide sequences with a type IA topoisomerase under conditions such that one strand, but not both strands, of one or both ends of a first ds nucleotide sequence are covalently linked by the topoisomerase. Compositions for performing such methods, and compositions generated from such methods also are provided, as are kits containing components useful for conveniently practicing the methods.

Abstract: The present invention provides nucleic acid molecules comprising one or more nucleic acid sequences encoding a polypeptide having a detectable activity. The present invention also provides methods of joining such nucleic acid molecules to nucleic acid molecules to be assayed for promoter activity. The present invention also relates to methods of preparing fusion proteins comprising a polypeptide of interest and a polypeptide having a detectable activity.

Abstract: The disclosure relates generally to genetic manipulation of stem and primary cells and to reprogramming of somatic cells, more specifically, human cells. In particular, compositions and methods are disclosed for the generation and maintenance of such engineered cells.

Abstract: The present invention provides nucleic acid molecules comprising one or more nucleic acid sequences encoding a polypeptide having a detectable activity. The present invention also provides methods of joining such nucleic acid molecules to nucleic acid molecules to be assayed for promoter activity. The present invention also relates to methods of preparing fusion proteins comprising a polypeptide of interest and a polypeptide having a detectable activity.

Abstract: The present invention relates to compositions and methods relating to development of modified inteins comprising one or more topoisomerase recognition sequences and the corresponding topoisomerase proteins and/or one or more recombination sites and the corresponding recombination proteins systems for use in affinity-based protein expression systems.

Abstract: Embodiments are directed to devices and methods for processing, cultivating or otherwise manipulating cell cultures which may be disposed on a flat or substantially flat surface such as cell culture substrate material. Devices and methods are disclosed for dividing a cell culture layer into divided portions, including isolated divided portions, that may then be transferred from the cell culture to a new location. For some embodiments, the divided portions may be transferred to a new cell culture support substrate in order to continue to grow and cultivate the cell line.

Abstract: The disclosure relates generally to stem cell biology and more specifically to genetic manipulation of stem cells. Methods and compositions using recombinational cloning techniques are disclosed which allow the construction and insertion of complex genetic constructs into embryonic and adult stem cells and progenitor cells. The methods disclosed will allow the harvesting of adult stem cells pre-engineered with integration sites to facilitate early passage genetic modification.

Abstract: The present invention provides compositions and methods for recombinational cloning. The compositions vectors having multiple recombination sites and/or multiple topoisomerase recognition sities. The methods premit the simultaeous cloning of two or more different nucleic acid molecules. In some embodiments the molecules are fused together while in other embodiments the molecules are inserted into distinct sites in a vector. The invention also generally provides for linking or joining through recombination a number of molecules and/or compounds (e.g., chemical compounds, drugs, proteins or peptides, lipids, nucleic acids, carbohydrates, etc.) which may be the same or different.

Abstract: A method of generating a double stranded (ds) recombinant nucleic acid molecule covalently linked in both strands by contacting two or more ds nucleotide sequences with a topoisomerase under conditions such that both termini of at least one end of a first ds nucleotide sequence are covalently linked by the topoisomerase to both termini of at least one end of a second ds nucleotide sequence is provided. Also provided is a method for generating a ds recombinant nucleic acid molecule covalently linked in one strand, by contacting two or more ds nucleotide sequences with a type IA topoisomerase under conditions such that one strand, but not both strands, of one or both ends of a first ds nucleotide sequence are covalently linked by the topoisomerase. Compositions for performing such methods, and compositions generated from such methods also are provided, as are kits containing components useful for conveniently practicing the methods.

Abstract: A method of generating a double stranded (ds) recombinant nucleic acid molecule covalently linked in both strands by contacting two or more ds nucleotide sequences with a topoisomerase under conditions such that both termini of at least one end of a first ds nucleotide sequence are covalently linked by the topoisomerase to both termini of at least one end of a second ds nucleotide sequence is provided. Also provided is a method for generating a ds recombinant nucleic acid molecule covalently linked in one strand, by contacting two or more ds nucleotide sequences with a type IA topoisomerase under conditions such that one strand, but not both strands, of one or both ends of a first ds nucleotide sequence are covalently linked by the topoisomerase. Compositions for performing such methods, and compositions generated from such methods also are provided, as are kits containing components useful for conveniently practicing the methods.

Abstract: The present invention provides compositions, methods, and kits for covalently linking nucleic acid molecules. The methods include a strand invasion step, and the compositions and kits are useful for performing such methods. For example, a method of covalently linking double stranded (ds) nucleic acid molecules can include contacting a first ds nucleic acid molecule, which has a topoisomerase linked to a 3? terminus of one end and has a single stranded 5? overhang at the same end, with a second ds nucleic acid molecule having a blunt end, such that the 5? overhang can hybridize to a complementary sequence of the blunt end of the second nucleic acid molecule, and the topoisomerase can covalently link the ds nucleic acid molecules. The methods are simpler and more efficient than previous methods for covalently linking nucleic acid sequences, and the compositions and kits facilitate practicing the methods, including methods of directionally linking two or more ds nucleic acid molecules.

Abstract: The present invention provides nucleic acid molecules comprising one or more nucleic acid sequences encoding a polypeptide having a detectable activity. The present invention also provides methods of joining such nucleic acid molecules to nucleic acid molecules to be assayed for promoter activity. The present invention also relates to methods of preparing fusion proteins comprising a polypeptide of interest and a polypeptide having a detectable activity.