This disclosure provides methods for monitoring an immune response. Methods comprise linking a polynucleotide sequence encoding a heavy chain variable region and a polynucleotide sequence encoding a light chain variable region from a single lymphocyte from a biological sample obtained before an immune response and linking a polynucleotide sequence encoding a heavy chain variable region and a polynucleotide sequence encoding a light chain variable region from a single lymphocyte from a biological sample obtained during or after an immune response. Methods further comprise performing high-throughput sequencing of the linked (paired) sequences from before the immune response and from during or after the immune response, and comparing the resulting sequence reads.

A bioinformatics process which provides an improved means to detect a JAK-STAT3 cellular signaling pathway in a subject, such as a human, based on the expression levels of at least three unique target genes of the JAK-STAT3 cellular signaling pathway measured in a sample. The invention includes an apparatus comprising a digital processor configured to perform such a method, a non-transitory storage medium storing instructions that are executable by a digital processing device to perform such a method, and a computer program comprising program code means for causing a digital processing device to perform such a method. Kits are also provided for measuring expression levels of unique sets of JAK-STAT3 cellular signaling pathway target genes.

The invention relates to a method for producing a ribonucleic acid (RNA) molecule composition comprising n different RNA molecule species, the method comprising a step of RNA in vitro transcription of a mixture of m different deoxyribonucleic acid (DNA) molecule species in a single reaction vessel in parallel, i.e. simultaneously, and a step of obtaining the RNA molecule composition. Also provided is the RNA composition provided by the inventive method and a pharmaceutical composition comprising the same as well as a pharmaceutical container. Moreover, the invention provides the RNA composition and the pharmaceutical composition for use as medicament.

The invention provides efficient methods for combining single-substitution libraries of nucleic acids that span and encode proteins of interest and for selecting resultant mutant proteins after expression which have improved properties or characteristics. Specifically, the methods comprising synthesizing a single substitution library for each of a plurality of domains of a protein; expressing separately each member of each single substitution library as a pre-candidate protein; selecting members of each single substitution library which encode pre-candidate proteins which exhibit an improvement in the one or more predetermined characteristics to form a selected library; shuffling members or the selected libraries in a PCR to produce a combinatorial shuffled library; expressing members of the shuffled library as candidate proteins; and selecting mutant proteins which have improved properties or characteristics.

Strategies, systems, methods, reagents, and kits for phage-assisted continuous evolution are provided herein. These include strategies, systems, methods, reagents, and kits allowing for stringency modulation to evolve weakly active or inactive biomolecule variants, negative selection of undesired properties, and/or positive selection of desired properties.

Strategies, systems, methods, reagents, and kits for phage-assisted continuous evolution are provided herein. These include strategies, systems, methods, reagents, and kits allowing for stringency modulation to evolve weakly active or inactive biomolecule variants, negative selection of undesired properties, and/or positive selection of desired properties.

A system of a recombinant bacteriophage library and a target of interest complex, wherein the recombinant bacteriophage peptide library includes a plurality of peptides expressed on the surface of recombinant bacteriophages wherein each recombinant bacteriophage includes (a) a pill protein; wherein each pill protein includes (b) a peptide or polypeptide involved in an intermolecular interaction, which differs by at least one amino acid from other peptides or polypeptides in the library; and (c) a modified protease cleavage site proximal to the peptide, wherein the modified protease cleavage site is the same in each bacteriophage, the modified cleavage site having a reduced binding affinity to a protease, as compared to a non-modified cleavage site, and wherein the target of interest complex includes a protease, a flexible linker attached to the protease, and a target of interest attached to the flexible linker, wherein the target of interest participates in an intermolecular interaction.

A system of a recombinant bacteriophage library and a target of interest complex, wherein the recombinant bacteriophage peptide library includes a plurality of peptides expressed on the surface of recombinant bacteriophages wherein each recombinant bacteriophage includes (a) a pill protein; wherein each pill protein includes (b) a peptide or polypeptide involved in an intermolecular interaction, which differs by at least one amino acid from other peptides or polypeptides in the library; and (c) a modified protease cleavage site proximal to the peptide, wherein the modified protease cleavage site is the same in each bacteriophage, the modified cleavage site having a reduced binding affinity to a protease, as compared to a non-modified cleavage site, and wherein the target of interest complex includes a protease, a flexible linker attached to the protease, and a target of interest attached to the flexible linker, wherein the target of interest participates in an intermolecular interaction.

Methods and compositions are provided for producing libraries of soluble random polypeptides. In the methods, the fraction of hydrophilic residues in the polypeptide is controlled so as to maintain the solubility of the polypeptide constructs.

Methods and compositions are provided for producing libraries of soluble random polypeptides. In the methods, the fraction of hydrophilic residues in the polypeptide is controlled so as to maintain the solubility of the polypeptide constructs.