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Team:Heidelberg/Templates/Del week13 overview
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==Amplification and purification of Del Genes from DSM-39== | ==Amplification and purification of Del Genes from DSM-39== | ||
| - | ===Goals=== | + | ====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. | 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==== | ||
{| class="wikitable" | {| class="wikitable" | ||
Revision as of 18:12, 4 October 2013
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 |  | |
| Primer FS_02 and FS_24 |  | |||
| DelA-F | Primer FS_02 and FS_05 | | ||
| DelA-G | Primer FS_02 and FS_26 |  | ||
| Primer FS_02 and FS_23 | | |||
| DelE | Primer FS_04 and FS_24 | | ||
| DelE-G | Primer FS_04 and FS_09 | | ||
| DelF-G | Primer FS_06 and FS_07 | | ||
| Primer FS_20 and FS_07 | | |||
| Primer FS_20 and FS_23 | | |||
| Primer FS_20 and FS_11_short | | |||
| Primer FS_21 and FS_07 | | |||
| Primer FS_21 and FS_11_short | | |||
| Primer FS_21 and FS_23 | | |||
| Primer FS_20 and FS_09 | | |||
| Primer FS_21 and FS_09 | | |||
| DelG | Primer FS_08 and FS_11_short | | ||
| Primer FS_08 and FS_23 | | |||
| DelO DelP | DelO-P | Primer FS_22 and FS_13 | | |
| Primer FS_22 and FS_13_short | | |||
| Re-PCRs | ||||
|---|---|---|---|---|
| Gene(s) | Fragment | Primer combination | Successful? | |
| DelG | DelG | Primer FS_10 and FS_23 | | |
| DelO DelP | DelO-P | Primer FS_22 and FS_13 | | |
| 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 | |
