TCR Rapid Assembly for Functional Testing (TCRAFT)
(Kit #
1000000271
)
Depositing Lab: Michael Birnbaum
The TCRAFT kit can be used to efficiently assemble paired αβ T-cell receptor (TCR) libraries from sequences. TCRAFT plasmids are available in a pre-pooled format (TCRAFT Pooled Library #1000000264) and individually here. This kit contains 48 vectors encoding TCR β variable (TRBV) alleles and the TCR α constant (TRAC) region, 45 vectors encoding TCR α variable (TRAV) alleles and the TCR β constant (TRBC) region, and a destination pHIV vector for the completed TCR library. These vectors are designed to be pooled but can be used individually to construct specific TCRs. To build a TCR library, users generate two oligo pools with the script linked in the protocol and perform three simple Golden Gate assembly reactions. The resulting TCR library can be expressed in cell lines or primary cells for functional screening to decode TCR specificity.
This kit will be sent as bacterial glycerol stocks in 96-well plate format.
Original Publication
Scalable TCR synthesis and screening enables antigen reactivity mapping in vitiligo. Gaglione SA, Mukkamala RS, Krishna C, Smith BE, Wadsworth II MH, Jelinsky SA, Perez CR, Schmidt-Hong L, Katz EL, Gellatly KJ, Ali LR, Shen J, Holec PV, Zhao QH, Chan AO , Xu EJK, Kravarik KM, Guzova JA, Dobson CS, Singh H, Garber M, Dougan M, Dougan SK, Harris JE, Winkler A, Birnbaum ME. bioRxiv 2025.04.24.649726. doi: 10.1101/2025.04.24.649726. bioRxiv 2025.04.24.649726 (Link opens in a new window)
Description
TCR Rapid Assembly for Functional Testing (TCRAFT) enables efficient assembly of paired αβ T-cell receptor (TCR) libraries from sequences by accurately linking TCR variable region genes to CDR3α/β-containing oligos through three Golden Gate reactions. TCRAFT plasmids are available in a pre-pooled format (TCRAFT Pooled Library #1000000264) and individually here. This kit contains:
- 48 plasmids encoding distinct TRBV alleles and the TRAC region
- 45 plasmids encoding distinct TRAV alleles and the TRBC region
- Destination pHIV plasmid for the completed TCR library
These plasmids are designed to be used in pools according to the TCRAFT protocol, but they can also be used to construct individual TCRs by selecting specific TRAV- and TRBV-encoding plasmids. To build a TCR library, users generate two oligo pools with the script linked in the protocol and perform three simple Golden Gate assembly reactions. The resulting TCR library can be expressed in cell lines or primary cells for functional screening to decode TCR specificity.
Figure 1: Human TCR library assembly.
How to Cite this Kit
These plasmids were created by your colleagues. Please acknowledge the Principal Investigator, cite the article in which they were created, and include Addgene in the Materials and Methods of your future publications.
For your Materials and Methods section:
"The TCRAFT Kit was a gift from Michael Birnbaum (Addgene kit #1000000271)."
For your Reference section:
Scalable TCR synthesis and screening enables antigen reactivity mapping in vitiligo. Gaglione SA, Mukkamala RS, Krishna C, Smith BE, Wadsworth II MH, Jelinsky SA, Perez CR, Schmidt-Hong L, Katz EL, Gellatly KJ, Ali LR, Shen J, Holec PV, Zhao QH, Chan AO , Xu EJK, Kravarik KM, Guzova JA, Dobson CS, Singh H, Garber M, Dougan M, Dougan SK, Harris JE, Winkler A, Birnbaum ME. bioRxiv 2025.04.24.649726. doi: 10.1101/2025.04.24.649726. bioRxiv 2025.04.24.649726 (Link opens in a new window)
TCR Rapid Assembly for Functional Testing (TCRAFT) - #1000000271
- Resistance Color Key
Each circle corresponds to a specific antibiotic resistance in the kit plate map wells.
- Inventory
Searchable and sortable table of all plasmids in kit. The Well column lists the plasmid well location in its plate. The Plasmid column links to a plasmid's individual web page.
- Kit Plate Map
96-well plate map for plasmid layout. Hovering over a well reveals the plasmid name, while clicking on a well opens the plasmid page.
Resistance Color Key
| Chloramphenicol | |
| Kanamycin | |
| Ampicillin |
Inventory
| Well | Plasmid | Resistance |
|---|---|---|
| A / 1 | TRBV2 |
|
| A / 2 | TRBV3-1 |
|
| A / 3 | TRBV4-1 |
|
| A / 4 | TRBV4-2 |
|
| A / 5 | TRBV4-3 |
|
| A / 6 | TRBV5-1 |
|
| A / 7 | TRBV5-4 |
|
| A / 8 | TRBV5-5 |
|
| A / 9 | TRBV5-6 |
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| A / 10 | TRBV5-8 |
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| A / 11 | TRBV6-1 |
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| A / 12 | TRBV6-2 |
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| B / 1 | TRBV6-3 |
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| B / 2 | TRBV6-4 |
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| B / 3 | TRBV6-5 |
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| B / 4 | TRBV6-6 |
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| B / 5 | TRBV6-8 |
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| B / 6 | TRBV6-9 |
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| B / 7 | TRBV7-2 |
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| B / 8 | TRBV7-3 |
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| B / 9 | TRBV7-4 |
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| B / 10 | TRBV7-6 |
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| B / 11 | TRBV7-7 |
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| B / 12 | TRBV7-8 |
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| C / 1 | TRBV7-9 |
|
| C / 2 | TRBV9 |
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| C / 3 | TRBV10-1 |
|
| C / 4 | TRBV10-2 |
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| C / 5 | TRBV10-3 |
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| C / 6 | TRBV11-1 |
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| C / 7 | TRBV11-2 |
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| C / 8 | TRBV11-3 |
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| C / 9 | TRBV12-3 |
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| C / 10 | TRBV12-4 |
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| C / 11 | TRBV12-5 |
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| C / 12 | TRBV13 |
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| D / 1 | TRBV14 |
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| D / 2 | TRBV15 |
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| D / 3 | TRBV16 |
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| D / 4 | TRBV18 |
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| D / 5 | TRBV19 |
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| D / 6 | TRBV20-1 |
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| D / 7 | TRBV24-1 |
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| D / 8 | TRBV25-1 |
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| D / 9 | TRBV27 |
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| D / 10 | TRBV28 |
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| D / 11 | TRBV29-1 |
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| D / 12 | TRBV30 |
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| E / 1 | TRAV1-1 |
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| E / 2 | TRAV1-2 |
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| E / 3 | TRAV2 |
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| E / 4 | TRAV3 |
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| E / 5 | TRAV4 |
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| E / 6 | TRAV5 |
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| E / 7 | TRAV6 |
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| E / 8 | TRAV7 |
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| E / 9 | TRAV8-1 |
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| E / 10 | TRAV8-2 |
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| E / 11 | TRAV8-3 |
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| E / 12 | TRAV8-4 |
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| F / 1 | TRAV8-6 |
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| F / 2 | TRAV9-1 |
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| F / 3 | TRAV9-2 |
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| F / 4 | TRAV10 |
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| F / 5 | TRAV12-1 |
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| F / 6 | TRAV12-2 |
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| F / 7 | TRAV12-3 |
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| F / 8 | TRAV13-1 |
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| F / 9 | TRAV13-2 |
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| F / 10 | TRAV14/DV4 |
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| F / 11 | TRAV16 |
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| F / 12 | TRAV17 |
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| G / 1 | TRAV18 |
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| G / 2 | TRAV19 |
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| G / 3 | TRAV20 |
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| G / 4 | TRAV21 |
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| G / 5 | TRAV22 |
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| G / 6 | TRAV23/DV6 |
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| G / 7 | TRAV24 |
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| G / 8 | TRAV25 |
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| G / 9 | TRAV26-1 |
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| G / 10 | TRAV26-2 |
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| G / 11 | TRAV27 |
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| G / 12 | TRAV29/DV5 |
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| H / 1 | TRAV30 |
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| H / 2 | TRAV34 |
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| H / 3 | TRAV35 |
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| H / 4 | TRAV36/DV7 |
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| H / 5 | TRAV38-1 |
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| H / 6 | TRAV38-2/DV8 |
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| H / 7 | TRAV39 |
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| H / 8 | TRAV40 |
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| H / 9 | TRAV41 |
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| H / 10 | pHIV-SapI-LacZ |
|