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Team:USyd-Australia/Notebook/Primers
From 2014.igem.org
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<tr><td> <h2>Primers</h2></td></tr> | <tr><td> <h2>Primers</h2></td></tr> | ||
<tr><td> | <tr><td> | ||
| - | <p>All primers were | + | <p>All primers were supplied by IDT.</p> |
<p>Primers were reconstituted in TE buffer to a stock concentration of 500μM. This is to minimise the number of freeze-thaw cycles that the primers undergo when being used. Dilutions were made to produce working primer stocks of 50μM or 10μM as required.</p> | <p>Primers were reconstituted in TE buffer to a stock concentration of 500μM. This is to minimise the number of freeze-thaw cycles that the primers undergo when being used. Dilutions were made to produce working primer stocks of 50μM or 10μM as required.</p> | ||
<p>To make 500μM stocks, Volume to add=(2x number of nMoles) μL | <p>To make 500μM stocks, Volume to add=(2x number of nMoles) μL | ||
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<tr><td><a name="iGEM1403"></a>iGEM1403</td><td>GTTCCACAGGGTAGCCAGCAGCATAGGATCCAAAGA</br>GTGCCACCTGACGTCTAAGAAACC</td><td>71.2C</td><td>primers for pSB prefix/suffix</td></tr> | <tr><td><a name="iGEM1403"></a>iGEM1403</td><td>GTTCCACAGGGTAGCCAGCAGCATAGGATCCAAAGA</br>GTGCCACCTGACGTCTAAGAAACC</td><td>71.2C</td><td>primers for pSB prefix/suffix</td></tr> | ||
<tr><td><a name="iGEM104"></a>iGEM1404</td><td>CCGAAAAGTGCCACCTGACGTCTAAGAAAGCTTCCC</br>CCTGATTCTGTGGATAACCGTATTACCG</td><td>70.3</td><td>primers for pSB prefix/suffix</td></tr> | <tr><td><a name="iGEM104"></a>iGEM1404</td><td>CCGAAAAGTGCCACCTGACGTCTAAGAAAGCTTCCC</br>CCTGATTCTGTGGATAACCGTATTACCG</td><td>70.3</td><td>primers for pSB prefix/suffix</td></tr> | ||
| - | <tr><td><a name="iGEM1405"></a>iGEM1405</td><td>GAGGTCGGACAGTATATTGAAACGTCAGGAGTCTAG</br> | + | <tr><td><a name="iGEM1405"></a>iGEM1405</td><td>GAGGTCGGACAGTATATTGAAACGTCAGGAGTCTAG</br>A<u>TACCGAGGGACAGACTACCAACTCACA</u></td><td> </td><td>primers for gene cassette circularisation (attC-aeBlue-aacC1)</td></tr> |
| - | <tr><td><a name="iGEM1406"></a>iGEM1406</td><td> GTATCTAGACTCCTGACGTTTCAATATACTGTCCG</br> | + | <tr><td><a name="iGEM1406"></a>iGEM1406</td><td> GTATCTAGACTCCTGACGTTTCAATATACTGTCCG</br>ACCTC<u>TTATTAGGTGGCGGTACTTGGGTCG</u></td><td> </td><td>primers for gene cassette circularisation (attC-aeBlue-aacC1)</td></tr> |
<tr><td><a name="iGEM1407"></a>iGEM1407 </td><td>CTATATCTATGATCTCGCAGTCTCC </td><td>53.8C </td><td> PCR primers over the junction between aacC1 (G block) and pSB1C3 vector, for validation of successful G block insertion by PCR screening</td></tr> | <tr><td><a name="iGEM1407"></a>iGEM1407 </td><td>CTATATCTATGATCTCGCAGTCTCC </td><td>53.8C </td><td> PCR primers over the junction between aacC1 (G block) and pSB1C3 vector, for validation of successful G block insertion by PCR screening</td></tr> | ||
<tr><td><a name="iGEM1408"></a>iGEM1408</td><td>GCCTTTTGCTCACATGTTCTTTC</td><td>55.2C</td><td>PCR primers over the junction between aacC1 (G block) and pSB1C3 vector, for validation of successful G block insertion by PCR screening</td></tr> | <tr><td><a name="iGEM1408"></a>iGEM1408</td><td>GCCTTTTGCTCACATGTTCTTTC</td><td>55.2C</td><td>PCR primers over the junction between aacC1 (G block) and pSB1C3 vector, for validation of successful G block insertion by PCR screening</td></tr> | ||
Latest revision as of 03:32, 18 October 2014
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Primers |
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All primers were supplied by IDT. Primers were reconstituted in TE buffer to a stock concentration of 500μM. This is to minimise the number of freeze-thaw cycles that the primers undergo when being used. Dilutions were made to produce working primer stocks of 50μM or 10μM as required. To make 500μM stocks, Volume to add=(2x number of nMoles) μL Below is a table of sequences of all primers used in the USyd-Australia 2014 iGEM project, accompanied by a brief description of its application |
| Name | Sequence (5'-3') | Theoretical Tm | Purpose |
|---|---|---|---|
| iGEM10 | AGCTTTCGCTAAGGATGATTTCTGGA | 58.2C | Screening and sequencing |
| iGEM16 | GAGTGCCACCTGACGTCTAAGAAACC | 60.9C | For BioBrick sequencing, amplification by PCR, or junction PCR. Alternative to iGEM VF2 primer |
| iGEM25 | CCCTGATTCTGTGGATAACCGTATTACCG | 60.0C | For BioBrick sequencing, amplification by PCR, or junction PCR. Alternative to iGEM VR primer |
| iGEM1401 | GGGAAGCTTTCTTAGACGTCAGGTGGCACTTTTCGG | 66.3C | pBBR replicon |
| iGEM1402 | TTTGGATCCTATGCTGCTGGCTACCCTGTGGAAC | 66.5C | pBBR replicon |
| iGEM1403 | GTTCCACAGGGTAGCCAGCAGCATAGGATCCAAAGAGTGCCACCTGACGTCTAAGAAACC | 71.2C | primers for pSB prefix/suffix |
| iGEM1404 | CCGAAAAGTGCCACCTGACGTCTAAGAAAGCTTCCCCCTGATTCTGTGGATAACCGTATTACCG | 70.3 | primers for pSB prefix/suffix |
| iGEM1405 | GAGGTCGGACAGTATATTGAAACGTCAGGAGTCTAGATACCGAGGGACAGACTACCAACTCACA | primers for gene cassette circularisation (attC-aeBlue-aacC1) | |
| iGEM1406 | GTATCTAGACTCCTGACGTTTCAATATACTGTCCGACCTCTTATTAGGTGGCGGTACTTGGGTCG | primers for gene cassette circularisation (attC-aeBlue-aacC1) | |
| iGEM1407 | CTATATCTATGATCTCGCAGTCTCC | 53.8C | PCR primers over the junction between aacC1 (G block) and pSB1C3 vector, for validation of successful G block insertion by PCR screening |
| iGEM1408 | GCCTTTTGCTCACATGTTCTTTC | 55.2C | PCR primers over the junction between aacC1 (G block) and pSB1C3 vector, for validation of successful G block insertion by PCR screening |
| iGEM1409 | GTTTTTTTGGATGGAGTGAAACGATGGCGATTG | 61.7C | Amplification of iGEM-IntI1 gBlock forward primer |
| iGEM1410 | TGACACCTTGCCCTTTTTTGCCG | 60.9C | Amplification of iGEM-IntI1 gBlock reverse primer |
| iGEM1411 | GTTTTTTTGGATGGAGTGAAACGATGGCGATTG | 61.7C | Gblock primers for IntI1 in pSAM-R |
| iGEM1412 | TGACACCTTGCCCTTTTTTG | 53.9C | Gblock primers for IntI1 in pSAM-R |
| iGEM1413 | GACATTGCCGTCACTGCGTC | 59.1C | Junction primers for IntI1 in pSAM-R |
| iGEM1414 | TGGTCCAGAACCTTGACCGAAC | 59.2C | Junction primers for IntI1 in pSAM-R |
| iGEM1415 | AACCGAGGATGCGAACCACTTC | 59.7C | Junction primers for IntI1 in pSAM-R |
| iGEM1416 | CCTTCAAACGTGCCGTAGAACAAGC | 60.4C | Junction primers for IntI1 in pSAM-R |
| iGEM1417 | AAATACTAGTAGCGGCCGCTGCAGTCCGGCAAAAAAG | 67.6C | Linearisation of pSB1C3, containing full BioBrick prefix and suffix |
| iGEM1418 | AAACTCTAGAAGCGGCCGCGAATTCCAGAAATCATCCTTAGC | 66.9C | Linearisation of pSB1C3, containing full BioBrick prefix and suffix |
| iGEM1419 | TTTGAATTCGCGGCCGCTTCTAGAGTACCGAGGGACAGAC | 68.9C | Amplification of aeBlue gBlock. Forward primer. |
| iGEM1420 | GGGCTGCAGCGGCCGCTACTAGTATTATTAGGTGG | 67.4C | Amplification of aeBlue gBlock. Reverse primer. |
| iGEM1421 | GGGAATTCAAATCTAGAGACACCATCG | 57.1C | Amplification of LacI-Plac-AttI gBlock. Forward primer |
| iGEM1422 | CCTGCAGTTTACTAGTTTTGCCTAACTTTGTTTTAG | 59.4C | Amplification of LacI-Plac-AttI gBlock. Reverse primer |
| iGEM1423 | CAGGCTTTACACTTTATGCTTCCG | 56.3C | lacI-Plac-attI RHJ-F right hand junction forward primer (use with iGEM25) |
| iGEM1424 | AATGTAATTCAGCTCCGCCATC | 55.5C | lacI-Plac-attI LHJ-R left hand junction reverse primer (use with iGEM16) |
| iGEM1425 | AAATCTAGATGGCGCGGCTTAACTCAGGTGTTAGGCTTAGGAGGCTCAAGTATGGGCAT | 70.7C | For making aacC1 gene cassettes from Tn1696. Use with iGEM1426. |
| iGEM1426 | AAAACTAGTGGCGTCGGCTTGGACGAATTGTTAGGCTTAGGTGGCGGTACTTGGGTC | 71.4C | For making aacC1 gene cassettes from Tn1696. Use with iGEM1425. |
| iGEM1427 | TTTTCTAGAGTTTGATGTTATGGAGCAGCAACG | 60.2C | For making aacC1 gene cassettes from Tn1696. Use with iGEM1428. Alternative to iGEM1424. |
| iGEM1428 | TTTACTAGTGCAAAAAGGCAGCAATTATGAGCC | 60.9C | For making aacC1 gene cassettes from Tn1696. Use with iGEM1427. Alternative to iGEM1426. |
| NVC71 | CTAAGAATCCATAGTCCAACTCC | 52.4C | aadB gene, rvs primer for junction screen |
| NVC92b | CACGCAAGACCTCAACCTTTTCC | 58.6C | aadB gene, rvs primer for junction screen |
| NVC158 | GATACCTTGTGCGGCTATGTCTG | 57.6C | pUS41/44, left of cloning site |
| NVC159 | GCCGCCTTGGGCCGGGTGATGTC | 69.2C | pUS41/44, right of cloning site |
With thanks to:
