Difference between revisions of "Part:BBa J3101:Design"
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=BioBricks= | =BioBricks= | ||
| − | The BioBricks designed for this part are not wild type. | + | The BioBricks designed for this part are not wild type. The following is the sequence with its BioBricks: |
| + | <font color='navy'>GAATTC</font color><font color='tan'>GCGGCCGC</font color>T<font color='orange'>TCTAGA</font color><font color='red'>-</font color><u>TTCGGGTGTCAACAATTGACCAAAATATTGATTTACAGCGTAATGCGCTTTCTAGTGCAAATTGTGACCGCATTTT</u><font color='blue'>G</font color><font color='gold'>ACTAGT</font color><font color='sky blue'>T</font color><font color='pink'>GCGGCCGC</font color><u>''C''</u><font color='blue'>TGCAG</font color> | ||
| + | |||
| + | There is no <font color='red'>G</font color> spacer between the <font color='orange'>XbaI</font color> and the <u>insert</u>. | ||
| + | |||
| + | The spacer between the <u>insert</u> and the <font color='gold'>SpeI</font color> site is a <font color='blue'>G</font color> instead of a T. | ||
| + | |||
| + | The spacer between the <font color='gold'>SpeI</font color> site and the <font color='pink'>NotI</font color> site is a <font color='sky blue'>T</font color> instead of an A. | ||
| + | |||
| + | The last <u>''C''</u> of the <font color='pink'>NotI</font color> site is not conserved with the initial C from the <font color='blue'>PstI</font color> site. | ||
| + | |||
| + | The BB suffix currently has this sequence for Not I and PstI: | ||
| + | <font color='pink'>GCGGCCG'''c'''</font color><font color='blue'>CTGCAG</font color> | ||
| + | |||
| + | But it should have been: | ||
| + | <font color='pink'>GCGGCCG</font color><font color='purple'>C</font color><font color='blue'>TGCAG</font color> | ||
===Source=== | ===Source=== | ||
Revision as of 19:17, 26 July 2006
Recombinational Enhancer (RE) for Hin/Hix inverting
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
Design Notes
The base pair change of a T from a C occurred during cloning but is located in the spacer region between the fis binding sites; therefore, we believe that it will still work.
The insertion is right before the biobrick ends and several bases after the distal fis binding site; therefore, we believe it will function correctly.
We chose this RE to use because after cloning it retained functional full biobricks and the mutations do not seem harmful to function. The other RE that were cloned retained the proper RE sequence but lost critical cut sites in the biobricks.
RE is cloned in plasmid pSB1A2.
BioBricks
The BioBricks designed for this part are not wild type. The following is the sequence with its BioBricks: GAATTCGCGGCCGCTTCTAGA-TTCGGGTGTCAACAATTGACCAAAATATTGATTTACAGCGTAATGCGCTTTCTAGTGCAAATTGTGACCGCATTTTGACTAGTTGCGGCCGCCTGCAG
There is no G spacer between the XbaI and the insert.
The spacer between the insert and the SpeI site is a G instead of a T.
The spacer between the SpeI site and the NotI site is a T instead of an A.
The last C of the NotI site is not conserved with the initial C from the PstI site.
The BB suffix currently has this sequence for Not I and PstI: GCGGCCGcCTGCAG
But it should have been: GCGGCCGCTGCAG
Source
References
Haykinson and Johnson. 1993. DNA looping and the helical repeat in vitro and in vivo: effect of HU protein and enhancer location on Hin invertasome assembly [http://www.pubmedcentral.gov/articlerender.fcgi?tool=pubmed&pubmedid=8508775]