p128Tol2-Olactb:Cas9-t2A-GFP, 4xU6:sgRNA

226834 Danio rerio Bacterial Expression

Plasmid Information

Purpose
Expression of b-actin driven Cas9-GFP and U6-driven 4 sgRNAs in zebrafish
Depositor
Bushra Raj
Article
Siniscalco et al Development. 2024 Nov 22:dev.203102. doi
Promoter
Olactb and U6
Available Since
Dec. 10, 2024
Availability
Available

Sequence Information

Length
15,621 bp
Topology
circular
Molecule Type
other DNA
View on AddGene.org

Features

CDS 7
Label Gene Product Position Strand
AmpR bla beta-lactamase 2477..3338 Fwd
DNA Sequence
ATGAGTATTCAACATTTCCGTGTCGCCCTTATTCCCTTTTTTGCGGCATTTTGCCTTCCTGTTTTTGCTCACCCAGAAACGCTGGTGAAAGTAAAAGATGCTGAAGATCAGTTGGGTGCACGAGTGGGTTACATCGAACTGGATCTCAACAGCGGTAAGATCCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAATGATGAGCACTTTTAAAGTTCTGCTATGTGGCGCGGTATTATCCCGTATTGACGCCGGGCAAGAGCAACTCGGTCGCCGCATACACTATTCTCAGAATGACTTGGTTGAGTACTCACCAGTCACAGAAAAGCATCTTACGGATGGCATGACAGTAAGAGAATTATGCAGTGCTGCCATAACCATGAGTGATAACACTGCGGCCAACTTACTTCTGACAACGATCGGAGGACCGAAGGAGCTAACCGCTTTTTTGCACAACATGGGGGATCATGTAACTCGCCTTGATCGTTGGGAACCGGAGCTGAATGAAGCCATACCAAACGACGAGCGTGACACCACGATGCCTGTAGCAATGGCAACAACGTTGCGCAAACTATTAACTGGCGAACTACTTACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAGGCGGATAAAGTTGCAGGACCACTTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTGATAAATCTGGAGCCGGTGAGCGTGGGTCTCGCGGTATCATTGCAGCACTGGGGCCAGATGGTAAGCCCTCCCGTATCGTAGTTATCTACACGACGGGGAGTCAGGCAACTATGGATGAACGAAATAGACAGATCGCTGAGATAGGTGCCTCACTGATTAAGCATTGGTAA
Protein Sequence
MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYIELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRWEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGASLIKHW
EGFP — the original enhanced GFP (Yang et al.,… 7920..8640 Rev
DNA Sequence
ATGGTGAGCAAGGGCGAAGAGCTGTTCACCGGGGTGGTGCCCATCCTGGTCGAGCTGGACGGCGACGTAAACGGCCACAAGTTCAGCGTGTCCGGCGAGGGCGAGGGCGATGCCACCTACGGCAAGCTGACCCTGAAGTTCATCTGCACCACCGGCAAGCTGCCCGTGCCCTGGCCCACCCTCGTGACCACCCTGACCTACGGCGTGCAGTGCTTCAGCCGCTACCCCGACCACATGAAGCAGCACGACTTCTTCAAGTCCGCCATGCCCGAAGGCTACGTCCAGGAGCGCACCATCTTCTTCAAGGACGACGGCAACTACAAGACCCGCGCCGAGGTGAAGTTCGAGGGCGACACCCTGGTGAACCGCATCGAGCTGAAGGGCATCGACTTCAAGGAGGACGGCAACATCCTGGGGCACAAGCTGGAGTACAACTACAACAGCCACAACGTCTATATCATGGCCGACAAGCAGAAGAACGGCATCAAGGTGAACTTCAAGATCCGCCACAACATCGAGGACGGCAGCGTGCAGCTCGCCGACCACTACCAGCAGAACACCCCCATCGGCGACGGCCCCGTGCTGCTGCCCGACAACCACTACCTGAGCACCCAGTCCGCCCTGAGCAAAGACCCCAACGAGAAGCGCGATCACATGGTCCTGCTGGAGTTCGTGACCGCCGCCGGGATCACTCTCGGCATGGACGAGCTGTACAAGTAA
Protein Sequence
MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDGNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEFVTAAGITLGMDELYK
EGFP — the original enhanced GFP (Yang et al.,… 7923..8640 Rev
DNA Sequence
ATGGTGAGCAAGGGCGAAGAGCTGTTCACCGGGGTGGTGCCCATCCTGGTCGAGCTGGACGGCGACGTAAACGGCCACAAGTTCAGCGTGTCCGGCGAGGGCGAGGGCGATGCCACCTACGGCAAGCTGACCCTGAAGTTCATCTGCACCACCGGCAAGCTGCCCGTGCCCTGGCCCACCCTCGTGACCACCCTGACCTACGGCGTGCAGTGCTTCAGCCGCTACCCCGACCACATGAAGCAGCACGACTTCTTCAAGTCCGCCATGCCCGAAGGCTACGTCCAGGAGCGCACCATCTTCTTCAAGGACGACGGCAACTACAAGACCCGCGCCGAGGTGAAGTTCGAGGGCGACACCCTGGTGAACCGCATCGAGCTGAAGGGCATCGACTTCAAGGAGGACGGCAACATCCTGGGGCACAAGCTGGAGTACAACTACAACAGCCACAACGTCTATATCATGGCCGACAAGCAGAAGAACGGCATCAAGGTGAACTTCAAGATCCGCCACAACATCGAGGACGGCAGCGTGCAGCTCGCCGACCACTACCAGCAGAACACCCCCATCGGCGACGGCCCCGTGCTGCTGCCCGACAACCACTACCTGAGCACCCAGTCCGCCCTGAGCAAAGACCCCAACGAGAAGCGCGATCACATGGTCCTGCTGGAGTTCGTGACCGCCGCCGGGATCACTCTCGGCATGGACGAGCTGTACAAG
Protein Sequence
MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDGNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEFVTAAGITLGMDELYK
T2A — 2A peptide from Thosea asigna virus cap… 8646..8700 Rev
DNA Sequence
GAGGGCAGAGGCAGTCTGCTGACATGCGGTGACGTGGAAGAGAATCCCGGCCCT
Protein Sequence
EGRGSLLTCGDVEENPGP
SV40 NLS — nuclear localization signal of SV40 (si… 8715..8736 Rev
DNA Sequence
CCTAAGAAGAAGAGAAAGGTG
Protein Sequence
PKKKRKV
Cas9 — Cas9 (Csn1) endonuclease from the Strep… 8751..12852 Rev
DNA Sequence
ATGGATAAGAAGTATAGCATCGGCCTGGATATTGGAACTAACTCCGTGGGTTGGGCAGTGATTACAGACGACTACAAGGTCCCTAGCAAGAAATTTAAGGTGCTGGGTAACACCGACAGGCACAGCATCAAGAAAAATCTGATTGGAGCCCTGCTGTTCGGTTCTGGAGAGACTGCCGAAGCAACACGCCTGAAAAGAACAGCAAGAAGGCGCTATACCAGAAGGAAGAATAGAATCTGTTACCTGCAGGAGATTTTCTCTAACGAAATGGCTAAGGTGGACGATTCATTCTTTCATAGGCTGGAGGAAAGTTTCCTGGTCGAGGAAGATAAGAAACACGAGCGCCATCCTATCTTTGGAAACATTGTGGACGAGGTCGCCTATCACGAAAAATACCCAACCATCTATCATCTGCGCAAGAAACTGGCTGACTCTACTGATAAAGCCGACCTGAGACTGATCTATCTGGCTCTGGCCCACATGATTAAGTTCAGGGGTCATTTTCTGATCGAGGGCGATCTGAACCCCGACAATTCCGATGTGGACAAGCTGTTCATCCAGCTGGTCCAGATTTACAATCAGCTGTTTGAGGAAAACCCTATTAATGCTTCCAGAGTGGACGCAAAAGCTATCCTGTCAGCCAGGCTGTCCAAGTCACGCAGACTGGAGAACCTGATTGCACAGCTGCCCGGAGAAAAGAGGAACGGTCTGTTTGGAAATCTGATCGCTCTGAGTCTGGGCCTGACTCCTAACTTCAAAAGCAATTTTGATCTGGCTGAGGACGCCAAACTGCAGCTGTCAAAGGACACATATGACGATGACCTGGATAACCTGCTGGCACAGATCGGAGATCAGTACGCTGACCTGTTCCTGGCTGCCAAAAATCTGTCCGACGCAATCCTGCTGTCAGATATTCTGAGAGTGAACAGCGAGATTACAAAAGCACCTCTGAGTGCCAGCATGATCAAGAGATATGACGAGCACCATCAGGATCTGACCCTGCTGAAGGCTCTGGTCAGGCAGCAGCTGCCAGAGAAGTACAAGGAAATTTTCTTTGATCAGTCCAAGAACGGCTACGCCGGTTATATCGACGGAGGCGCATCACAGGAGGAATTCTACAAGTTTATCAAACCTATTCTGGAGAAGATGGACGGAACTGAGGAACTGCTGGTGAAACTGAATAGAGAGGACCTGCTGAGGAAGCAGCGCACATTTGATAACGGTTCCATCCCACACCAGATTCATCTGGGAGAGCTGCACGCTATCCTGAGGCGCCAGGAAGACTTCTACCCCTTTCTGAAAGATAACCGCGAGAAGATCGAAAAAATTCTGACCTTCAGAATCCCTTACTATGTGGGTCCACTGGCTCGCGGAAACAGCAGATTTGCCTGGATGACTCGCAAATCCGAGGAAACCATTACTCCTTGGAACTTCGAGGAAGTGGTCGATAAGGGCGCCTCTGCACAGTCCTTCATCGAGAGAATGACTAATTTTGACAAAAACCTGCCCAATGAGAAAGTGCTGCCTAAGCACTCCCTGCTGTACGAGTATTTCACTGTCTATAACGAACTGACAAAGGTGAAATACGTCACCGAGGGCATGAGAAAGCCAGCCTTCCTGTCAGGAGAGCAGAAGAAAGCAATCGTGGATCTGCTGTTTAAAACCAACAGGAAAGTGACTGTCAAGCAGCTGAAGGAGGACTACTTCAAGAAAATTGAATGCTTCGATTCCGTGGAGATCAGCGGAGTCGAAGACAGATTTAACGCAAGCCTGGGCGCTTACCACGATCTGCTGAAGATCATTAAGGATAAAGACTTCCTGGACAACGAGGAAAATGAGGATATCCTGGAAGACATTGTGCTGACACTGACCCTGTTTGAGGACAGAGGAATGATCGAGGAAAGACTGAAAACCTATGCTCATCTGTTCGATGACAAGGTGATGAAACAGCTGAAGAGAAGGCGCTACACTGGCTGGGGTAGACTGAGCAGGAAGCTGATCAACGGCATTAGGGATAAACAGTCAGGAAAGACAATCCTGGACTTTCTGAAAAGTGATGGCTTCGCCAACCGCAATTTTATGCAGCTGATTCACGATGACAGTCTGACCTTCAAAGAGGACATCCAGAAGGCTCAGGTGTCTGGACAGGGCCACTCCCTGCATGAGCAGATTGCAAACCTGGCTGGAAGCCCAGCCATCAAGAAAGGCATTCTGCAGACAGTGAAAATCGTCGATGAGCTGGTGAAAGTCATGGGCCATAAGCCCGAAAACATCGTGATTGAGATGGCTCGCGAAAATCAGACAACCCAGAAGGGTCAGAAGAACAGTAGAGAGAGGATGAAAAGAATCGAGGAAGGCATTAAGGAGCTGGGTAGCCAGATCCTGAAAGAGCACCCAGTGGAAAACACACAGCTGCAGAATGAGAAGCTGTATCTGTACTATCTGCAGAATGGAAGAGATATGTACGTGGACCAGGAGCTGGATATTAACAGGCTGTCTGATTACGACGTGGATCATATCGTCCCCCAGAGTTTCATCAAAGATGACAGCATTGACAACAAGGTGCTGACCAGGTCCGACAAAAACAGAGGAAAATCAGATAATGTCCCTAGTGAGGAAGTGGTCAAGAAAATGAAGAACTACTGGAGACAGCTGCTGAATGCCAAACTGATCACTCAGAGGAAGTTTGATAACCTGACAAAAGCAGAGCGCGGTGGACTGTCAGAACTGGACAAAGCTGGATTCATCAAGAGGCAGCTGGTGGAAACACGCCAGATCACTAAACACGTCGCACAGATTCTGGATAGTCGCATGAACACAAAGTACGATGAGAATGACAAACTGATCAGAGAAGTGAAGGTCATTACCCTGAAGAGTAAACTGGTCAGCGACTTTAGGAAAGATTTCCAGTTTTATAAGGTCCGCGAGATTAACAATTATCACCATGCCCATGACGCATACCTGAACGCCGTGGTCGGTACCGCACTGATCAAGAAATACCCAAAACTGGAGAGCGAATTCGTGTACGGAGACTATAAGGTGTACGATGTCAGAAAAATGATCGCCAAGTCCGAGCAGGAAATTGGAAAAGCTACTGCCAAGTATTTCTTTTACTCAAACATCATGAATTTCTTTAAGACAGAGATCACCCTGGCCAATGGAGAAATCCGCAAAAGGCCCCTGATTGAGACAAACGGAGAGACAGGCGAAATCGTGTGGGACAAAGGCAGAGATTTTGCAACCGTGAGGAAGGTCCTGAGCATGCCTCAAGTGAATATCGTCAAGAAAACTGAGGTGCAGACAGGCGGTTTCTCAAAAGAAAGTATTCTGCCAAAACGCAACTCTGATAAGCTGATCGCTAGAAAGAAAGACTGGGACCCTAAGAAGTATGGAGGCTTTGACTCTCCCACTGTGGCATACTCCGTCCTGGTGGTCGCTAAGGTGGAGAAGGGCAAAAGCAAGAAACTGAAATCTGTCAAGGAGCTGCTGGGTATCACAATTATGGAGAGAAGCTCTTTCGAGAAGAACCCAATCGATTTTCTGGAGGCCAAAGGTTATAAGGAAGTGAAGAAAGACCTGATCATTAAACTGCCCAAGTACAGTCTGTTTGAGCTGGAAAACGGCAGGAAACGCATGCTGGCAAGCGCTGGAGAGCTGCAGAAAGGCAATGAACTGGCCCTGCCTTCTAAGTACGTGAACTTCCTGTATCTGGCAAGCCACTACGAGAAGCTGAAAGGATCTCCAGAGGATAACGAACAGAAACAGCTGTTTGTGGAGCAGCACAAGCATTATCTGGACGAGATCATTGAACAGATTAGCGAGTTCTCTAAAAGAGTGATCCTGGCCGACGCAAATCTGGATAAGGTCCTGTCTGCTTACAACAAACACAGAGATAAGCCCATCAGGGAGCAGGCCGAAAATATCATTCATCTGTTCACTCTGACAAACCTGGGCGCACCTGCAGCTTTCAAGTACTTCGACACTACAATCGATAGAAAGAGGTACACCTCCACTAAGGAGGTGCTGGACGCTACACTGATCCATCAGAGTATTACCGGCCTGTACGAAACAAGGATTGACCTGTCTCAGCTGGGTGGCGAC
Protein Sequence
MDKKYSIGLDIGTNSVGWAVITDDYKVPSKKFKVLGNTDRHSIKKNLIGALLFGSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESFLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLADSTDKADLRLIYLALAHMIKFRGHFLIEGDLNPDNSDVDKLFIQLVQIYNQLFEENPINASRVDAKAILSARLSKSRRLENLIAQLPGEKRNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQIGDQYADLFLAAKNLSDAILLSDILRVNSEITKAPLSASMIKRYDEHHQDLTLLKALVRQQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLRKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGNSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYFTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDSVEISGVEDRFNASLGAYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDRGMIEERLKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQLIHDDSLTFKEDIQKAQVSGQGHSLHEQIANLAGSPAIKKGILQTVKIVDELVKVMGHKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYLYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFIKDDSIDNKVLTRSDKNRGKSDNVPSEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHVAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLNAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTEITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGITIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNELALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLDATLIHQSITGLYETRIDLSQLGGD
SV40 NLS — nuclear localization signal of SV40 (si… 12858..12879 Rev
DNA Sequence
CCTAAGAAGAAGAGAAAGGTG
Protein Sequence
PKKKRKV
RBS 1
Label Gene Product Position Strand
Shine-Dalgarno sequence — — 8830..8839 Fwd
DNA Sequence
ACCTCCTTA
polyA_signal 2
Label Gene Product Position Strand
SV40 poly(A) signal — — 5262..5397 Fwd
DNA Sequence
AACTTGTTTATTGCAGCTTATAATGGTTACAAATAAAGCAATAGCATCACAAATTTCACAAATAAAGCATTTTTTTCACTGCATTCTAGTTGTGGTTTGTCCAAACTCATCAATGTATCTTATCATGTCTGGATC
SV40 poly(A) signal — — 7676..7811 Fwd
DNA Sequence
AACTTGTTTATTGCAGCTTATAATGGTTACAAATAAAGCAATAGCATCACAAATTTCACAAATAAAGCATTTTTTTCACTGCATTCTAGTTGTGGTTTGTCCAAACTCATCAATGTATCTTATCATGTCTGGATC
promoter 7
Label Gene Product Position Strand
T7 promoter — — 1216..1235 Rev
DNA Sequence
TAATACGACTCACTATAGG
AmpR promoter bla — 2372..2477 Fwd
DNA Sequence
CGCGGAACCCCTATTTGTTTATTTTTCTAAATACATTCAAATATGTATCCGCTCATGAGACAATAACCCTGATAAATGCTTCAATAATATTGAAAAAGGAAGAGT
lac promoter — — 4420..4451 Fwd
DNA Sequence
TTTACACTTTATGCTTCCGGCTCGTATGTTG
T3 promoter — — 4519..4538 Fwd
DNA Sequence
AATTAACCCTCACTAAAGG
T3 promoter — — 7865..7884 Fwd
DNA Sequence
AATTAACCCTCACTAAAGG
T7 promoter — — 12932..12951 Rev
DNA Sequence
TAATACGACTCACTATAGG
T3 promoter — — 13002..13021 Fwd
DNA Sequence
AATTAACCCTCACTAAAGG
protein_bind 6
Label Gene Product Position Strand
attB4 mutant version of attB — 1236..1257 Rev
DNA Sequence
CAACTTTGTATAGAAAAGTTG
CAP binding site — — 4384..4406 Fwd
DNA Sequence
TAATGTGAGTTAGCTCACTCAT
lac operator — — 4458..4475 Fwd
DNA Sequence
TTGTGAGCGGATAACAA
attB3 mutant version of attB — 7593..7614 Fwd
DNA Sequence
CAACTATGTATAATAAAGTTG
attB2 mutant version of attB — 7812..7837 Fwd
DNA Sequence
CCCACTTTGTACAAGAAAGCTGGGT
attB1 mutant version of attB — 12978..13003 Rev
DNA Sequence
TCAAGTTTGTACAAAAAAGCAGGCT
rep_origin 1
Label Gene Product Position Strand
ori — — 3508..4097 Fwd
DNA Sequence
TTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGCAAACAAAAAAACCACCGCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTTTTTCCGAAGGTAACTGGCTTCAGCAGAGCGCAGATACCAAATACTGTTCTTCTAGTGTAGCCGTAGTTAGGCCACCACTTCAAGAACTCTGTAGCACCGCCTACATACCTCGCTCTGCTAATCCTGTTACCAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGACGATAGTTACCGGATAAGGCGCAGCGGTCGGGCTGAACGGGGGGTTCGTGCACACAGCCCAGCTTGGAGCGAACGACCTACACCGAACTGAGATACCTACAGCGTGAGCTATGAGAAAGCGCCACGCTTCCCGAAGGGAGAAAGGCGGACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGGAGAGCGCACGAGGGAGCTTCCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTTCGCCACCTCTGACTTGAGCGTCGATTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATGGAAA