Team:Wageningen UR/Journal/week13

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= week 13: 23 july - 27 july =
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= week 13: 23 july - 29 july =
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Not much work was done this week because the team spent four days on going to a three day synthetic biology conference in Munich. We learned a lot there and had a great time!
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== Office work ==
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== Lab work ==
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''' CCMV '''
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''27th July'' (Hugo)
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<br/>
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Another PCR repeat of 18-07's PCR. Purified PCR products using Fermentas PCR Purification Kit then ran another PCR reaction (repeat of 19-07).
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<br/>
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<br/>
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''28th July'' (Jeroen)
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* Digested IPTG inducible promotor with RBS brick: Spe1 and Pst1
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* Digested CCMV pJET fragment with Xba1 and Pst1
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* after 30 minutes used a PCR purification for digestion inactivation (Pst1 is not heat inactivatable)
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* Ligation 1 hour
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* Transformation of DH5alpha cells
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''30th July'' (Jeroen)
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* 5 colonies picked for colony PCR per plate
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'''PLRV'''
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* RNA isolation of potato leaf tissue
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* Reverse transcriptase reaction to obtain cDNA
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* PCR on cDNA using coat protein primers
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The figure below shows the purity of isolated RNA. We isolated RNA of different potato cultivars.
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[[File:rna2.jpg|frame|center|''Figure 1: isolated RNA PLRV'']]
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[[File:table1.jpg|frame|center|''Figure 2: ...'']]
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We performed Reverse Transciptase reactions followed by PCR. We used the Revert AID H Minus Reverse Transcription Kit (Fermentas).
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[[File:rt-pcr.jpg|frame|center|''Figure 3: ...'']]
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Agarose gel of PCR products. Lanes 1 till 4 show the PLRV Coat Protein (CP) bands with the expected size of 627 bp. Lanes 6 till 9 show the PLRV Coat Protein plus Read Through part (CP_RT) with the expected size of 2154 bp.
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== Lab work ==
 
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RNA isolation of potato leaf tissue
 
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Reverse transcriptase reaction to obtain cDNA
 
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PCR on cDNA using coat protein primers
 
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Written by: Wouter
 
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Latest revision as of 03:26, 27 September 2012

week 13: 23 july - 29 july

Not much work was done this week because the team spent four days on going to a three day synthetic biology conference in Munich. We learned a lot there and had a great time!

Lab work

CCMV

27th July (Hugo)
Another PCR repeat of 18-07's PCR. Purified PCR products using Fermentas PCR Purification Kit then ran another PCR reaction (repeat of 19-07).

28th July (Jeroen)

  • Digested IPTG inducible promotor with RBS brick: Spe1 and Pst1
  • Digested CCMV pJET fragment with Xba1 and Pst1
  • after 30 minutes used a PCR purification for digestion inactivation (Pst1 is not heat inactivatable)
  • Ligation 1 hour
  • Transformation of DH5alpha cells

30th July (Jeroen)

  • 5 colonies picked for colony PCR per plate


PLRV

  • RNA isolation of potato leaf tissue
  • Reverse transcriptase reaction to obtain cDNA
  • PCR on cDNA using coat protein primers


The figure below shows the purity of isolated RNA. We isolated RNA of different potato cultivars.

Figure 1: isolated RNA PLRV
Figure 2: ...


We performed Reverse Transciptase reactions followed by PCR. We used the Revert AID H Minus Reverse Transcription Kit (Fermentas).

Figure 3: ...

Agarose gel of PCR products. Lanes 1 till 4 show the PLRV Coat Protein (CP) bands with the expected size of 627 bp. Lanes 6 till 9 show the PLRV Coat Protein plus Read Through part (CP_RT) with the expected size of 2154 bp.




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