Team:Heidelberg/Templates/Del week13 overview

From 2013.igem.org

Amplification and purification of Del Genes from DSM-39

Goals

As our the amplifications of of the sequence region DelF-G did not progress last week, we decided to focus on this region in week 13. All combinations of initial and newly ordered primers will be tried with various PCR protocols. Furthermore, we tried to get higher amounts of the fragments for which protocols have already been established in order to have enough of each product to carry out suitable test restriction digests.

Results

PCRs from D.acidovorans DSM-39
Gene(s) Fragment Primer combination Successful?
DelA
DelB
DelC
DelD
DelE
DelF
DelG
DelA-E Primer FS_02 and FS_03
Heidelberg Yes check.svg.png
Primer FS_02 and FS_24
Heidelberg Tilde.png
DelA-F Primer FS_02 and FS_05
Heidelberg Yes check.svg.png
DelA-G Primer FS_02 and FS_26
Heidelberg Red x.svg.png
Primer FS_02 and FS_23
Heidelberg Red x.svg.png
DelE Primer FS_04 and FS_24
Heidelberg Red x.svg.png
DelE-G Primer FS_04 and FS_09
Heidelberg Red x.svg.png
DelF-G Primer FS_06 and FS_07
Heidelberg Red x.svg.png
Primer FS_20 and FS_07
Heidelberg Tilde.png
Primer FS_20 and FS_23
Heidelberg Red x.svg.png
Primer FS_20 and FS_11_short
Heidelberg Red x.svg.png
Primer FS_21 and FS_07
Heidelberg Red x.svg.png
Primer FS_21 and FS_11_short
Heidelberg Red x.svg.png
Primer FS_21 and FS_23
Heidelberg Red x.svg.png
Primer FS_20 and FS_09
Heidelberg Red x.svg.png
Primer FS_21 and FS_09
Heidelberg Red x.svg.png
DelG Primer FS_08 and FS_11_short
Heidelberg Red x.svg.png
Primer FS_08 and FS_23
Heidelberg Yes check.svg.png
DelO
DelP
DelO-P Primer FS_22 and FS_13
Heidelberg Red x.svg.png
Primer FS_22 and FS_13_short
Heidelberg Tilde.png



Re-PCRs
Gene(s) Fragment Primer combination Successful?
DelG DelG Primer FS_10 and FS_23
Heidelberg Yes check.svg.png
DelO
DelP
DelO-P Primer FS_22 and FS_13
Heidelberg Tilde.png



Test restriction digests of PCR amplified fragments
Fragment Primer Digestion enzyme Expected bands [bp]
DelA-E Primer FS_02 and FS_03 EcoRI-HF 3054, 2260
DelA-F Primer FS_02 and FS_05 EcoRI-HF 4624, 4354, 2260
DelE-G Primer FS_04 and FS_07 PvuI-HF 6187, 4917
DelF-G Primer FS_20 and FS_07 XmaI 4307, 879
Primer FS_20 and FS_07 ClaI 2420, 1519, 1247
DelG Primer FS_08 and FS_23 BglII 3291, 1936, 1303
DelO-P Primer FS_22 and FS_13_short EcoRI-HF 1883, 960