Team:Panama/28 September 2010
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==='''September 28'''=== | ==='''September 28'''=== | ||
- | '''Digestion of | + | '''Digestion of Rh1AB with Pst1''' |
- | Since the | + | Since the RhlAB gene is too big, we haven´t been able to complete the sequencing necessary for us to know if we're on the right track. We think that we can test it with a PstI digestion. We have checked the expected cuts with the NEBcutter (of NEB)and it is possible to but the Rhlab gene with Pst1. After this we have to do: |
-Electrophoresis | -Electrophoresis | ||
- | Clone | + | Clone Rh1AB into the plasmid. |
-This week | -This week | ||
- | PCR to show the | + | PCR to show the Rh1AB gene. |
- | + | ||
- | + | ||
The gene was purified with the Roche Kit: High Pure PCR Product Purification Kit | The gene was purified with the Roche Kit: High Pure PCR Product Purification Kit | ||
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- | Run a gel to see the disgestion of the gene and to verify the cut sites of | + | Run a gel to see the disgestion of the gene and to verify the cut sites of PstI |
[[Image:Cuadro28SEPTIEMBRE.JPG|350px|thumb|left|alt text]] | [[Image:Cuadro28SEPTIEMBRE.JPG|350px|thumb|left|alt text]] | ||
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+ | [[Image:GEL28SEPTIEMBRE1i.jpg|300px|thumb|left|alt text]] | ||
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Cloning Protocol of the pGEM-T Vector System | Cloning Protocol of the pGEM-T Vector System | ||
- | [[Image: | + | [[Image:Cuadro28setiembre.jpg|500px|thumb|left|alt text]] |
Latest revision as of 03:33, 28 October 2010
September 28
Digestion of Rh1AB with Pst1
Since the RhlAB gene is too big, we haven´t been able to complete the sequencing necessary for us to know if we're on the right track. We think that we can test it with a PstI digestion. We have checked the expected cuts with the NEBcutter (of NEB)and it is possible to but the Rhlab gene with Pst1. After this we have to do:
-Electrophoresis
Clone Rh1AB into the plasmid.
-This week
PCR to show the Rh1AB gene.
The gene was purified with the Roche Kit: High Pure PCR Product Purification Kit
The purification of the gene RH1ab was done because it is a requirement in order to be able to use with the pGEM-T Easy Vector system and to clone it into a vector to later do the mutagenesis
Run a gel to see the disgestion of the gene and to verify the cut sites of PstI
1) We don't see the expected bands, possibly because we aren't using a purified PCR product
2) We can't see the product. We will have to repeat the ligation
3) We observe the band at the expected size
Measurement of the concentration of the purification: 96.1 ug/ml
Cloning Protocol of the pGEM-T Vector System
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