2274 - Ongoing Supply of Molecular Biology Consumables

Ongoing supply of enzymes. Enzymes should be tested for exonuclease and endonuclease activity. All enzymes should be provided with the manufacturer's definition of a "unit" of enzyme e.g. "one U of enzyme is defined as the amount of enzyme required to digest xµg of DNA at 37°C in xx minutes in a total reaction volume of xxµl"

CPI require columns for use in packed column chromatography during the purification of in vitro transcribed mRNA.

CPI require AAV material as a control or standard for downstream and analytical assays such as ELISA and western blotting. This material would be used to check the validity of assays and act as a comparison to in-house generated samples. Further, the material would be used to develop and validate methods for different analyses prior to analysis of samples generated in-house. The material required would need to be of a high purity with the associated CoA for each item purchased.

CPI require the supply of ELISA kits for the detection of proteins during the production of different therapeutics such host cell protein contaminants, recombinant therapeutic proteins and AAV particles. A range of kits are required that target and confirm the presence of specific protein targets in samples. For AAV analysis, different serotypes exist, and specific kits allow the distinction between these types and confirm the formation of AAV particles. ELISA kits are required to be specific to the intended target and include the CoA/quality control certificates.

CPI require reporter assays as an efficient method to measure cell health during experiments. Assays allow of aspects such as product related toxicity and detect differences in cell growth. A range of kits are required that allow real-time measurements, compatibility with manual handling and/or automation. Reporter assays are required to provide high sensitivity, accurate, stable readouts and applicable across different cell types with the necessary CoA/quality control certificates.

CPI require the supply of transfection reagents to deliver nucleic acids in different formats (RNA/DNA) into cells for the testing or production of therapeutics.

dNTPs and NTPs will be required for use in multiple PCR applications; in vitro transcription; mRNA/saRNA synthesis and cloning. dNTPs and NTPs should be possible of being supplied in sodium or tris salt form and confirmed as RNase, DNase and protease free. The ability to supply alternative salt forms, such as lithium, would be advantageous. These dNTPs and NTPs will be used for a range of applications in both small scale and large scale, it would be beneficial if they could be supplied in custom bulk volumes.

CPI requires the synthesis of primers, DNA oligos, and gene synthesis services. CPI would require random primer sequences and the ability to buy custom synthesised primers for use in PCR/qPCR and would also require custom primer/ oligo synthesis, CPI will provide the sequence information for these primers to be synthesised. We would require primers to be produced at a selection of purities, such as HPLC and salt free. The supply of fluorescently labelled primer probes will also be required under this lot. For gene synthesis, CPI require a flexibility to synthesise a range of sequences varying in size (typically up to 4kb, with varying complexities) with use a variety of plasmid backbone options with a range of antibiotic resistance genes.

CPI would require consumables for the analysis of proteins and nucleic acids by gel electrophoresis. Consumables required include gel loading dyes; RNA, DNA and protein ladders and gel extraction kits. RNA and DNA ladders should span a range of sizes, up to 9kb. Concentrated gel running buffers such as 10XTAE, 10XTBE and MOPS is advantageous. The supply of other purification kits such as gel extraction kits and genomic DNA extraction kits are advantageous.

Concentrated buffers and nuclease free water for use with their associated enzymes during reactions are required. Buffers may be used where specified by the manufacturer in applications which include restriction reactions and in vitro transcription reactions. Buffers must be tested for exonuclease and endonuclease activity and a CoA must be available for each lot number. These buffers are likely to be used in both small scale and large-scale applications and so custom bulk volumes of buffers would be advantageous. GMP grade options are also advantageous.

CPI would require competent cells to be supplied for use in cloning/sub-cloning, protein expression and gDNA/cDNA library production.

CPI require the supply of RNA and DNA purification kits. RNA/DNA purification kits can be either gravity flow, vacuum assisted or spin columns and should cover small scale through to large scale. Multiple types of PCR kits would be required for use in development activities at CPI also, such as RT-PCR and RT-qPCR kits.

CPI require the supply of cell culture media to assist with the testing of therapeutic nucleic acids (DNA/RNA). Cell culture media and feeds are required, as either pre-made sterile solutions or as a powder.

CPI require supply of the RNA co-translational capping agent, cleancap. The supplier must be able to supply various types of therapeutically relevant cleancap analogues, such as AU and AG (in addition to the methylated form).