Team:NJU China/Project/Brain

From 2013.igem.org

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<a>Liver:</a>
<a>Liver:</a>
<span>For liver targeting, we need to first find a protein specifically recognize hepatic cells. Since Heptitis B virus can infect hepatic cells distinctively, and from recent study[1], we knew that HBV recognizes the hepatic cells via the interaction between the pre-S1 of the HBV envelop protein and NTCP receptor of the hepatic cells. We tried to engineer the pre-S1 from HBV envelope protein to the lamp 2b.</BR>
<span>For liver targeting, we need to first find a protein specifically recognize hepatic cells. Since Heptitis B virus can infect hepatic cells distinctively, and from recent study[1], we knew that HBV recognizes the hepatic cells via the interaction between the pre-S1 of the HBV envelop protein and NTCP receptor of the hepatic cells. We tried to engineer the pre-S1 from HBV envelope protein to the lamp 2b.</BR>

Revision as of 12:31, 26 September 2013

 

RESULTS

Brain

2013

Liver: For liver targeting, we need to first find a protein specifically recognize hepatic cells. Since Heptitis B virus can infect hepatic cells distinctively, and from recent study[1], we knew that HBV recognizes the hepatic cells via the interaction between the pre-S1 of the HBV envelop protein and NTCP receptor of the hepatic cells. We tried to engineer the pre-S1 from HBV envelope protein to the lamp 2b.
Therefore we cloned the pre-S1 into lamp 2b, and we choose pcDNA 3.1(+) as our backbone.

Results for liver targeting: To produce the exosomes that have pre-S1 on their surface for liver targeting, we first transfected the exosome-producing cells, HEK 293T cells, with the plasmid encoding the fusion protein of lamp 2b and pre-S1 peptide.

Liver

2013

siRNA screening (Failed) 14th: We extracted plasmids and cultured the 293t cells. 15th: We transfected plasmids into 293t cells. 16th: We collected cells and preserved it in Trizol. 17th: We extracted RNA, then did RT-PCR with the RNA. 18th: We did qPCR with the cDNA we got on 17th April. 21th: We reexamined the concentration of RNA, and redid qPCR.

siRNA screening (Failed) 27th: We cultured the 293t cells in a 12-well plate. 28th: We transfected 293t cells. 29th: We collected cells and extracted RNA from it. 30th: We did RT-qPCR with the RNA we extracted on 29th April. 1st: We did qPCR with the cDNA we got on 30th April.

T cell

2013

siRNA screening (Failed), examination whether siRNA are capsulated into exosomes (Success) 8th: We extracted plasmids of 3 kinds of siRNA. 9th: We cultured 293t cells in eight D10 dishes and a 12-well plate. 10th: We extracted 2 kinds of over-expression plasmids and examined the concentration of it. Then we transfect these plasmids into 293t cells respectively. 11th: We collected cells and exosomes from 8 D10 dish, and cells from 12-well plate. 12th: We extracted RNA from cells and exosomes. 13th: We did RT-PCR and qPCR.

August

2013

July

2013

June

2013