Team:WLC-Milwaukee/Parts
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Overview
The 2021 WLC-Milwaukee iGEM team contributed many functional parts to the registry this year. Our lacY and lacI biobricks were synthesized by IDT Technologies and inserted into pSB1C3. The lacY and lacI biobricks were transformed into E. coli DH5a. Our bla constructs were synthesized de novo by TwistBioscience and are in their pTwistChlor Medium copy plasmid. These parts were transformed into E. coli MC4100.
We improved the part BBa_Q04121 by using the open reading frame of the lacI gene and adding its original promoter. We also added regulatory elements and a promoter and ribosome binding site for any gene that is inserted downstream of our lacI biobrick.
We improved the part BBa_I763015 by adding a 40 base spacer from the original genome, thereby including its natural ribosome binding site. Additionally, the 40 base spacer ensures that RNA polymerase has ample room to bind the lacY promoter. Furthermore, the ORF of lacY is integrated into all our devices, demonstrating its improvement.
The sequences of the lacY and lacI biobricks can be found in our bla constructs. This demonstrates their functionality as they are integral to the color change induced by the bla construct in the presence of nitrocefin.
Basic Parts
| Part Number | Type | Description |
|---|---|---|
| BBa_K4062000 | Regulatory | lacI Promoter |
| BBa_K4062001 | Coding | lacI ORF |
| BBa_K4062002 | Regulatory | CAP Site |
| BBa_K4062003 | Regulatory | lacP |
| BBa_K4062004 | Regulatory | lacO |
| BBa_K4062005 | RBS | lacZ RBS |
| BBa_K4062006 | Coding | bla ORF |
| BBa_K4062007 | RBS | lacY RBS |
| BBa_K4062008 | Coding | lacY ORF |
| BBa_K4062009 | Regulatory | lacI Promoter, -10 deletion |
| BBa_K4062010 | Regulatory | lacP, -10 deletion |
| BBa_K4062011 | Regulatory | CAP site, deletion |
| BBa_K4062012 | Coding | lacI ORF, ORF stop mutation |
| BBa_K4062017 | Regulatory | lacO, mutation |
| BBa_K4062018 | RBS | lacZ RBS, mutation |
Composite Parts
| Part Number | Type | Description |
|---|---|---|
| BBa_K4062014 | Composite | lacI + Regulatory components + lacZ RBS |
| BBa_K4062015 | Composite | lacY + RBS |
| BBa_K4062016 | Device | Lactose regulated b-lactamase expression |
| BBa_K4062023 | Device | Lactose regulated b-lactamase expression, lacI promomoter inactivated |
| BBa_K4062024 | Device | Lactose regulated b-lactamase expression, lacP deletion |
| BBa_K4062025 | Device | Lactose regulated b-lactamase expression, CAP mutation |
| BBa_K4062026 | Device | Lactose regulated b-lactamase expression, lacI ORF mutation |
| BBa_K4062027 | Device | Lactose regulated b-lactamase expression, lacO mutation |
| BBa_K4062028 | Device | Lactose regulated b-lactamase expression, lacZ RBS mutation |
| BBa_K4062030 | Composite | lacI + Regulatory components + lacZ RBS, lacI promoter deletion |
| BBa_K4062031 | Composite | lacI + Regulatory components + lacZ RBS, lacP deletion |
| BBa_K4062032 | Composite | lacI + Regulatory components + lacZ RBS, CAP site deletion |
| BBa_K4062033 | Composite | lacI + Regulatory components + lacZ RBS, lacI ORF mutation |
| BBa_K4062034 | Composite | lacI + Regulatory components + lacZ RBS, lacO mutation |
| BBa_K4062035 | Composite | lacI + Regulatory components + lacZ RBS, lacZ RBS mutation |
