OPTIMIZATION OF THE PRODUCTION OF PLASMID PCDNA3.1(+) UTR_SPIKE FOR COVID-19 MRNA VACCINE THROUGH VARIATION OF HARVESTING TIME AND CULTIVATION MEDIA
Plasmid is a genetic material that is often used in the process of molecular cloning. One of its applications in the health sector is in the development of COVID-19 mRNA vaccines. The production of plasmid in host cells is significantly influenced by the harvesting time and specific composition i...
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| Format: | Final Project |
| Language: | Indonesia |
| Online Access: | https://digilib.itb.ac.id/gdl/view/87640 |
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| Institution: | Institut Teknologi Bandung |
| Language: | Indonesia |
| Summary: | Plasmid is a genetic material that is often used in the process of molecular cloning. One
of its applications in the health sector is in the development of COVID-19 mRNA
vaccines. The production of plasmid in host cells is significantly influenced by the
harvesting time and specific composition in the cultivation media used. This study aims
to determine the optimal harvesting time for E. coli TOP10 and the effect of variation in
components in the cultivation media on the production performance of
pcDNA3.1(+)_UTR_Spike, a COVID-19 mRNA vaccine. Transformed E. coli TOP10
host cell colonies with the pcDNA3.1(+)_UTR_Spike plasmid were first inoculated from
glycerol stock to solid LB (Lysogeny Broth) media, and then a single colony was
inoculated into various treated liquid media. The media used were LB, TB (Terrific Broth),
and TBGL (TB + glycerol 46 g/L). Plasmid isolation was performed at 12 hours and 24
hours after the preparation of the working culture. The results showed that the volumetric
yield of plasmid between the isolates at 12 hours and 24 hours was not significantly
different. The 12-hour isolate from the TB media showed a volumetric yield ~1.8X lower
than that from the LB media, while the TBGL media showed a volumetric yield ~2.7X
lower compared to the LB media. It can be concluded that the harvesting of E. coli TOP10
for the production of pcDNA3.1(+)_UTR_Spike should ideally be done at 12 hours postpreparation
of the working culture. The components of tryptone 10g/L, yeast extract 5g/L,
and NaCl 10g/L found in the LB (Miller) media were able to produce a better production
performance of pcDNA3.1(+)_UTR_Spike compared to the TB and TBGL media, with
an average plasmid volumetric yield of 12.52 ?g/mL.
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