Split Selectable Markers

▲Nate (left), Albert (center), Menghan (right). Photography by Charles Camarda.

Split selectable markers. Selectable markers are widely used in transgenesis and genome editing for selecting engineered cells with a desired genotype but the variety of markers is limited. Split selectable markers are created by dividing fluorescent proteins or antibiotic resistance proteins into several segments fused to protein splicing elements called “inteins” can be separately co-segregated with different transgenic vectors, and rejoin via protein trans-splicing to reconstitute a full-length marker protein in host cells receiving all intended vectors. In a work led by lab manager/RA Nathaniel Jillette and PhD candidate graduate student Menghan Du, we demonstrated utilities of split selectable markers in lentiviral transgenesis and CRISPR/Cas-mediated genome editing.

▲Selection achieved by intein-split resistance (intres) genes.

Publication:

Jillette, N., Du, M., Zhu, J.J., Cardoz, P., Cheng, A.W. (2019) Split Selectable Markers. Nature Communications 10:4968. doi: 10.1038/s41467-019-12891-2 PMID: 31672965 PMCID: PMC6823381

Press release:

“Improving transgene research with split selectable markers” by Mark Wanner

Grant support:

R01HG009900, P30CA034196.

Plasmid Listing. Genbank/sequence files available through Addgene links.

Plasmid # Plasmid Name Addgene Link
3 pLX-Hygro(1-89)-NpuDnaE(N)-IRES-TagBFP2 http://www.addgene.org/122730
4 pLX-NpuDnaE(C)-Hygro(90-341)-IRES-mCherry http://www.addgene.org/122731
5 pLX-Hygro(1-200)-SspDnaB(N)-IRES-TagBFP2 http://www.addgene.org/122732
6 pLX-SspDnaB(C)-Hygro(201-341)-IRES-mCherry http://www.addgene.org/122733
7 pLX-Hygro(1-52)-NpuDnaE(N)-IRES-TagBFP2 http://www.addgene.org/122734
8 pLX-NpuDnaE(C)-Hygro(53-341)-IRES-mCherry http://www.addgene.org/122735
9 pLX-Hygro(1-240)-NpuDnaE(N)-IRES-TagBFP2 http://www.addgene.org/122736
10 pLX-NpuDnaE(C)-Hygro(241-341)-IRES-mCherry http://www.addgene.org/122737
11 pLX-Hygro(1-292)-NpuDnaE(N)-IRES-TagBFP2 http://www.addgene.org/122738
12 pLX-NpuDnaE(C)-Hygro(293-341)-IRES-mCherry http://www.addgene.org/122739
13 pLX-Blast(1-102)-NpuDnaE(N)-IRES-TagBFP2 http://www.addgene.org/122740
14 pLX-NpuDnaE(C)-Blast(103-140)-IRES-mCherry http://www.addgene.org/122741
17 pLX-Puro(1-119)-NpuDnaE(N)-IRES-TagBFP2 http://www.addgene.org/122742
18 pLX-NpuDnaE(C)-Puro(insCys;120-199)-IRES-mCherry http://www.addgene.org/122743
19 pLX-Puro(1-100)-SspDnaB(N-S0)-IRES-TagBFP2 http://www.addgene.org/122744
20 pLX-SspDnaB(C-S0)-Puro(101-199)-IRES-mCherry http://www.addgene.org/122745
21 pLX-Neo(1-133)-NpuDnaE(N)-IRES-TagBFP2 http://www.addgene.org/122746
22 pLX-NpuDnaE(C)-Neo(134-267)-IRES-mCherry http://www.addgene.org/122747
23 pLX-Neo(1-194)-NpuDnaE(N)-IRES-TagBFP2 http://www.addgene.org/122748
24 pLX-NpuDnaE(C)-Neo(195-267)-IRES-mCherry http://www.addgene.org/122749
25 pLX-NpuDnaE(C)_Hygro(53-89)-NpuDnaE(N)-IRES-GFP http://www.addgene.org/122750
26 pLX-NpuDnaE(C)_Hygro(53-239)-NpuDnaE(N)-IRES-GFP http://www.addgene.org/122751
27 pCR8-BsaI->ccdbCam<-BsaI-NpuDnaE(N) http://www.addgene.org/122752
28 pCR8-NpuDnaE(C)_BsaI->ccdbCam<-BsaI http://www.addgene.org/122753
31 pCR8-BsaI->ccdbCam<-BsaI-SspDnaB(N-S0) http://www.addgene.org/122754
32 pCR8-SspDnaB(C-S0)_BsaI->ccdbCam<-BsaI http://www.addgene.org/122755
33 pLX-mScarlet(1-46)-NpuDnaE(N)_LZA-IRES-TagBFP2 http://www.addgene.org/122756
34 pLX-LZB_NpuDnaE(C)-mScarlet(insCys;47-232)-IRES-TagBFP2 http://www.addgene.org/122757
35 pLX-mScarlet(1-48)-NpuDnaE(N)_LZA-IRES-TagBFP2 http://www.addgene.org/122758
36 pLX-LZB_NpuDnaE(C)-mScarlet(insCys;49-232)-IRES-GFP http://www.addgene.org/122759
37 pLX-mScarlet(1-51)-NpuDnaE(N)_LZA -IRES-TagBFP2 http://www.addgene.org/122760
38 pLX-LZB_NpuDnaE(C)-mScarlet(insCys;52-232)-IRES-GFP http://www.addgene.org/122761
39 pLX-mScarlet(1-75)-NpuDnaE(N)_LZA-IRES-TagBFP2 http://www.addgene.org/122762
40 pLX-LZB_NpuDnaE(C)-mScarlet(insCys;76-232)-IRES-GFP http://www.addgene.org/122763
41 pLX-mScarlet(1-122)-NpuDnaE(N)_LZA-IRES-TagBFP2 http://www.addgene.org/122764
42 pLX-LZB_NpuDnaE(C)-mScarlet(insCys;123-232)-IRES-GFP http://www.addgene.org/122765
43 pLX-mScarlet(1-140)-NpuDnaE(N)_LZA-IRES-TagBFP2 http://www.addgene.org/122766
44 pLX-LZB_NpuDnaE(C)-mScarlet(insCys;141-232)-IRES-GFP http://www.addgene.org/122767
45 pLX-mScarlet(1-163)-NpuDnaE(N)_LZA-IRES-TagBFP2 http://www.addgene.org/122768
46 pLX-LZB_NpuDnaE(C)-mScarlet(insCys;164-232)-IRES-GFP http://www.addgene.org/122769
47 pCR8-TagBFP2 http://www.addgene.org/122770
48 pCR8-mCherry http://www.addgene.org/122771
49 pLX-DEST-IRES-Hygro(1-89)-NpuDnaE(N) http://www.addgene.org/122772
50 pLX-DEST-IRES-NpuDnaE(C)-Hygro(90-341) http://www.addgene.org/122773
51 pLX-[TagBFP2]-IRES-Hygro(1-89)-NpuDnaE(N) http://www.addgene.org/122774
52 pLX-[mCherry]-IRES-NpuDnaE(C)-Hygro(90-341) http://www.addgene.org/122775
53 pLX-DEST-IRES-Puro(1-119)-NpuDnaE(N) http://www.addgene.org/122776
54 pLX-DEST-IRES-NpuDnaE(C)-Puro(120-199) http://www.addgene.org/122777
55 pLX-[TagBFP2]-IRES-Puro(1-119)-NpuDnaE(N) http://www.addgene.org/122778
56 pLX-[mCherry]-IRES-NpuDnaE(C)-Puro(120-199) http://www.addgene.org/122779
57 pLX-DEST-IRES-Neo(1-194)-NpuDnaE(N) http://www.addgene.org/122780
58 pLX-DEST-IRES-NpuDnaE(C)-Neo(195-267) http://www.addgene.org/122781
59 pLX-[TagBFP2]-IRES-Neo(1-194)-NpuDnaE(N) http://www.addgene.org/122782
60 pLX-[mCherry]-IRES-NpuDnaE(C)-Neo(195-267) http://www.addgene.org/122783
64 pLX-Hygro(1-69)-NpuDnaE(N)-IRES-TagBFP2 http://www.addgene.org/122784
65 pLX-NpuDnaE(C)-Hygro(^C;70-341)-IRES-mCherry http://www.addgene.org/122785
66 pLX-Hygro(1-131)-NpuDnaE(N)-IRES-TagBFP2 http://www.addgene.org/122786
67 pLX-NpuDnaE(C)-Hygro(^C;132-341)-IRES-mCherry http://www.addgene.org/122787
68 pLX-Hygro(1-171)-NpuDnaE(N)-IRES-TagBFP2 http://www.addgene.org/122788
69 pLX-NpuDnaE(C)-Hygro(^C;172-341)-IRES-mCherry http://www.addgene.org/122789
70 pLX-Hygro(1-218)-NpuDnaE(N)-IRES-TagBFP2 http://www.addgene.org/122790
71 pLX-NpuDnaE(C)-Hygro(^C;219-341)-IRES-mCherry http://www.addgene.org/122791
72 pLX-Hygro(1-259)-NpuDnaE(N)-IRES-TagBFP2 http://www.addgene.org/122792
73 pLX-NpuDnaE(C)-Hygro(^C;260-341)-IRES-mCherry http://www.addgene.org/122793
74 pLX-Hygro(1-277)-NpuDnaE(N)-IRES-TagBFP2 http://www.addgene.org/122794
75 pLX-NpuDnaE(C)-Hygro(^C; 278-341)-IRES-mCherry http://www.addgene.org/122795
76 pLX-Puro(1-32)-NpuDnaE(N)-IRES-TagBFP2 http://www.addgene.org/122796
77 pLX-NpuDnaE(C)-Puro(^C;33-199)-IRES-mCherry http://www.addgene.org/122797
78 pLX-Puro(1-84)-NpuDnaE(N)-IRES-TagBFP2 http://www.addgene.org/122798
79 pLX-NpuDnaE(C)-Puro(^C;85-199)-IRES-mCherry http://www.addgene.org/122799
80 pLX-Puro(1-137)-NpuDnaE(N)-IRES-TagBFP2 http://www.addgene.org/122800
81 pLX-NpuDnaE(C)-Puro(^C;138-199)-IRES-mCherry http://www.addgene.org/122801
82 pLX-Puro(1-158)-NpuDnaE(N)-IRES-TagBFP2 http://www.addgene.org/122802
83 pLX-NpuDnaE(C)-Puro(^C;159-199)-IRES-mCherry http://www.addgene.org/122803
84 pLX-Puro(1-180)-NpuDnaE(N)-IRES-TagBFP2 http://www.addgene.org/122804
85 pLX-NpuDnaE(C)-Puro(^C;181-199)-IRES-mCherry http://www.addgene.org/122805
86 pLX-Blast(1-58)-NpuDnaE(N)-IRES-TagBFP2 http://www.addgene.org/122806
87 pLX-NpuDnaE(C)-Blast(59-140)-IRES-mCherry http://www.addgene.org/122807
88 pLX-NpuDnaE(C)-HygroBA-SspDnaB(N-S0)-IRES-EGFP http://www.addgene.org/122808
89 pLX-SspDnaB(C-S0)-Hygro(201-341)-IRES-mCherry http://www.addgene.org/122809
90 pLX-NpuDnaE(C)-Hygro(90-200)-SspDnaB(N-S0)-IRES-EGFP http://www.addgene.org/122810
91 pLX-Hygro(1-200)-SspDnaB(N-S0)-IRES-TagBFP2 http://www.addgene.org/122811
92 pLX-SspDnaB(C-S0)-Hygro(201-240)-NpuDnaE(N)-IRES-EGFP http://www.addgene.org/122812
93 pLX-SspDnaB(C-S0)-Hygro(201-292)-NpuDnaE(N)-IRES-EGFP http://www.addgene.org/122813
94 pLX-DEST-IRES-TagBFP2 http://www.addgene.org/122814
95 pLX-DEST-IRES-EGFP http://www.addgene.org/122815
96 pLX-DEST-IRES-mCherry http://www.addgene.org/122816
97 pLX-Hygro-IRES-TagBFP2 http://www.addgene.org/122817
98 pLX-Hygro-IRES-mCherry http://www.addgene.org/122818
99 pLX-Puro-IRES-TagBFP2 http://www.addgene.org/122819
100 pLX-Puro-IRES-mCherry http://www.addgene.org/122820
101 pLX-Hygro-IRES-EGFP http://www.addgene.org/122821
102 pLX-NLS_GFP-IRES-Hygro http://www.addgene.org/122822
103 pLX-LifeAct_mCherry-IRES-Hygro http://www.addgene.org/122823
104 pLX-NLS_GFP-IRES-Hygro(1-89)-NpuDnaE(N) http://www.addgene.org/122824
105 pLX-LifeAct_mScarlet-IRES- NpuDnaE(C)-Hygro(90-341) http://www.addgene.org/122825
106 pX330-AAVS1 http://www.addgene.org/122826
107 pAAVS1-Nst-EF1aHygro2ArtTA3(-)_TetO-Blast-P2A-EGFP http://www.addgene.org/122827
108 pAAVS1-Nst-EF1aHygro2ArtTA3(-)_TetO-Blast-P2A-mScarlet http://www.addgene.org/122828
109 pAAVS1-Nst-EF1aHygro2ArtTA3(-)_TetO-Blast(1-102)_NpuDnaE(N)-P2A-EGFP http://www.addgene.org/122829
110 pAAVS1-Nst-EF1aHygro2ArtTA3(-)_TetO-NpuDnaE(C)_Blast(103-140)-P2A-mScarlet http://www.addgene.org/122830
111 pAAVS1-Nst-EF1aBlast2ArtTA3(-)_TetO-Hygro-P2A-NTR-E2A-EGFP http://www.addgene.org/122831
112 pAAVS1-Nst-EF1aBlast2ArtTA3(-)_TetO-Hygro-P2A-NTR-E2A-mCherry http://www.addgene.org/122832
113 pAAVS1-Nst-EF1aBlast2ArtTA3(-)_TetO- Hygro(1-89)-NpuDnaE(N)-P2A-NTR-E2A-EGFP http://www.addgene.org/122833
114 pAAVS1-Nst-EF1aBlast2ArtTA3(-)_TetO- NpuDnaE(C)-Hygro(90-341)-P2A-NTR-E2A-mCherry http://www.addgene.org/122834