cancer vaccine
A cancer vaccine, cancer antigen technology, applied in vaccines, DNA/RNA vaccination, cancer antigen components, etc., can solve the problems of activation, insertional mutagenesis, inhibition, etc.
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Examples
Embodiment 1
[0751] Example 1. Production of polynucleotides
[0752] According to the present disclosure, the manufacture of polynucleotides and parts or regions thereof can be achieved using the methods taught in the International Application WO2014 / 152027 entitled "Manufacturing Methods for Production of RNA Transcripts", the contents of which are incorporated by reference Incorporated into this article as a whole.
[0753] Purification methods may include those taught in International Applications WO2014 / 152030 and WO2014 / 152031, each of which is incorporated herein by reference in its entirety.
[0754] Methods of detection and characterization of polynucleotides can be performed as taught in WO2014 / 144039, which is hereby incorporated by reference in its entirety.
[0755] Characterization of polynucleotides of the present disclosure can be accomplished using a program selected from the group consisting of polynucleotide mapping, reverse transcriptase sequencing, charge distributi...
Embodiment 2
[0756] Embodiment 2 chimeric polynucleotide synthesis
[0757] introduction
[0758] According to the present disclosure, two regions or portions of a chimeric polynucleotide can be joined or linked using triphosphate chemistry.
[0759] According to this approach, a first region or portion of 100 nucleotides or less is chemically synthesized with a 5' monophosphate and a terminal 3' de-OH or blocked OH. If the region is longer than 80 nucleotides, it can be synthesized as two strands for ligation.
[0760] If the first region or portion is synthesized as a non-positionally modified region or portion using in vitro transcription (IVT), then conversion to the 5' monophosphate and subsequent capping of the 3' end can be followed.
[0761] Monophosphate protecting groups may be selected from any of those known in the art.
[0762] The second region or portion of the chimeric polynucleotide can be synthesized using chemical synthesis or IVT methods. IVT methods can include...
Embodiment 3
[0774] Example 3: PCR for cDNA Production
[0775] 2x KAPA HIFI manufactured by Kapa Biosystems (Woburn, MA) was used TM The HotStart ReadyMix performs the PCR procedure for the preparation of cDNA. This system includes 12.5 μl 2x KAPA ReadyMix; 0.75 μl forward primer (10 μM); 0.75 μl reverse primer (10 μM); 100 ng template cDNA; 2 0 diluted to 25.0 μl. Reaction conditions were 95°C for 5 minutes, and 25 cycles of 98°C for 20 seconds, then 58°C for 15 seconds, then 72°C for 45 seconds; then 72°C for 5 minutes, then 4°C until termination.
[0776] Use Invitrogen's PURELINK according to manufacturer's instructions (up to 5 μg) TM PCR Mini Kit (Carlsbad, CA) was used to clean up the reaction. Larger volume reactions will require purification using products with greater throughput. After cleanup, the cDNA was quantified using NANODROP™ and analyzed by agarose gel electrophoresis to confirm that the cDNA was of the expected size. The cDNA is then submitted for sequencing ...
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Description
Claims
Application Information
- IPC
- A61K9/10; A61K48/00; A61K39/00
- CPC
- A61K39/0011; A61K39/001114; A61K9/5123; A61P35/00; A61K2039/53; A61K2039/545; A61K2039/55588; A61K9/10
- Inventors
- 尼古拉斯·瓦利安特; 塔尔·扎克斯