pDIV495

177702 Synthetic Bacterial Expression

Plasmid Information

Purpose
Plasmid expressing optimized Cas9 and NAT marker and sgRNA targeting ADE2 locus in D. hansenii
Depositor
Uffe Mortensen
Article
Strucko et al Synth Biol (Oxf). 2021 Oct 12;6(1):ysab0
Tags
SV40
Promoter
RNR2p (Debaryomyces hansenii ), SRC1p (Debaryomyc…
Available Since
March 11, 2022
Availability
Available

Sequence Information

Length
12,088 bp
Topology
circular
Molecule Type
other DNA
View on AddGene.org

Features

CDS 6
Label Gene Product Position Strand
AmpR bla beta-lactamase 2690..3551 Rev
DNA Sequence
ATGAGTATTCAACATTTCCGTGTCGCCCTTATTCCCTTTTTTGCGGCATTTTGCCTTCCTGTTTTTGCTCACCCAGAAACGCTGGTGAAAGTAAAAGATGCTGAAGATCAGTTGGGTGCACGAGTGGGTTACATCGAACTGGATCTCAACAGCGGTAAGATCCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAATGATGAGCACTTTTAAAGTTCTGCTATGTGGCGCGGTATTATCCCGTATTGACGCCGGGCAAGAGCAACTCGGTCGCCGCATACACTATTCTCAGAATGACTTGGTTGAGTACTCACCAGTCACAGAAAAGCATCTTACGGATGGCATGACAGTAAGAGAATTATGCAGTGCTGCCATAACCATGAGTGATAACACTGCGGCCAACTTACTTCTGACAACGATCGGAGGACCGAAGGAGCTAACCGCTTTTTTGCACAACATGGGGGATCATGTAACTCGCCTTGATCGTTGGGAACCGGAGCTGAATGAAGCCATACCAAACGACGAGCGTGACACCACGATGCCTGTAGCGATGGCAACAACGTTGCGCAAACTATTAACTGGCGAACTACTTACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAGGCGGATAAAGTTGCAGGACCACTTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTGATAAATCCGGAGCCGGTGAGCGTGGTTCTCGCGGTATCATCGCAGCGCTGGGGCCAGATGGTAAGCCCTCCCGTATCGTAGTTATCTACACGACGGGGAGTCAGGCAACTATGGATGAACGAAATAGACAGATCGCTGAGATAGGTGCCTCACTGATTAAGCATTGGTAA
Protein Sequence
MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYIELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRWEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGASLIKHW
Cas9 — Cas9 (Csn1) endonuclease from the Strep… 6758..10862 Fwd
DNA Sequence
ATGGATAAAAAGTATAGTATTGGTTTAGATATTGGTACTAACTCTGTGGGTTGGGCAGTTATCACTGACGAATATAAAGTTCCATCAAAGAAATTTAAGGTGTTAGGTAACACTGACAGACACTCAATAAAAAAGAATCTTATCGGTGCTCTTTTGTTCGACTCCGGTGAAACTGCCGAGGCTACACGTTTAAAAAGAACAGCAAGAAGAAGATATACCCGTAGAAAAAATAGAATATGTTATTTACAAGAAATCTTTTCTAATGAAATGGCTAAAGTTGATGATTCCTTTTTCCATAGATTGGAAGAGTCATTTTTGGTTGAAGAAGACAAAAAGCATGAGAGACATCCAATCTTTGGGAATATAGTTGATGAAGTGGCTTACCATGAAAAATATCCTACCATTTATCATTTAAGAAAGAAATTGGTAGATTCAACTGATAAAGCTGACCTTAGATTAATCTATTTAGCACTTGCCCATATGATTAAATTTAGAGGTCATTTTTTGATTGAAGGTGATTTGAACCCAGATAATTCTGACGTGGATAAATTATTTATTCAATTAGTCCAAACCTACAACCAATTATTTGAGGAAAATCCAATTAATGCTAGTGGTGTCGATGCCAAAGCTATATTATCAGCCAGATTATCAAAATCTAGACGTTTGGAAAATTTGATTGCCCAATTGCCAGGAGAAAAAAAGAATGGATTATTTGGAAACTTGATCGCATTATCATTGGGTTTGACACCAAATTTTAAATCTAATTTTGATTTAGCTGAAGATGCTAAATTACAATTATCAAAAGACACCTATGACGACGATTTGGACAATTTACTTGCTCAAATTGGTGATCAATATGCAGATTTGTTCTTAGCTGCTAAAAACTTATCTGATGCTATTTTGTTGTCTGATATTTTGAGAGTGAACACAGAAATAACCAAAGCTCCATTATCAGCATCTATGATCAAACGTTATGATGAACACCATCAGGATTTGACTTTATTGAAAGCTTTGGTGAGACAACAATTGCCAGAGAAGTATAAAGAAATCTTTTTCGATCAATCTAAAAACGGGTATGCAGGTTATATTGATGGGGGTGCCTCCCAAGAGGAATTTTACAAATTTATAAAACCTATTTTAGAAAAGATGGATGGGACTGAGGAACTTTTGGTCAAATTGAACAGAGAAGATTTGTTACGTAAACAGAGAACTTTTGATAATGGTAGTATACCTCACCAAATTCATTTGGGTGAGTTGCATGCAATTTTAAGAAGACAAGAAGATTTTTATCCATTTTTAAAAGATAATAGAGAAAAAATCGAGAAAATTTTAACCTTTAGAATTCCATACTATGTTGGGCCTTTGGCTAGAGGTAATTCAAGATTTGCCTGGATGACACGTAAATCAGAAGAAACTATTACCCCTTGGAATTTTGAAGAGGTTGTTGATAAAGGAGCATCAGCACAGAGTTTTATTGAAAGAATGACCAATTTCGATAAAAACTTACCAAATGAAAAAGTTTTACCAAAACATTCCTTGTTATACGAATATTTTACTGTTTACAATGAACTTACAAAGGTTAAATATGTTACTGAAGGTATGCGTAAGCCAGCCTTTTTATCTGGAGAACAGAAAAAGGCAATAGTTGATTTATTGTTTAAAACAAATAGAAAAGTTACTGTTAAACAATTAAAAGAAGATTACTTTAAGAAAATTGAATGTTTTGATTCAGTTGAAATCAGTGGTGTTGAAGACAGATTTAATGCTAGTTTAGGAACTTACCATGATTTACTTAAAATTATCAAAGATAAAGATTTCTTGGATAACGAAGAAAATGAAGACATTTTAGAAGACATTGTTTTAACCTTAACTTTATTCGAAGATAGAGAGATGATTGAAGAACGTTTGAAGACTTATGCACATTTGTTTGACGATAAAGTGATGAAACAGTTGAAAAGAAGACGTTATACTGGATGGGGTAGATTGTCTCGTAAATTGATCAATGGAATTAGAGATAAACAAAGTGGTAAAACTATCTTGGACTTTTTGAAATCTGACGGATTTGCTAATAGAAATTTCATGCAATTGATCCACGACGATAGTTTGACATTTAAAGAAGACATCCAAAAGGCCCAAGTGAGTGGGCAAGGTGATTCATTACATGAACATATTGCAAATTTAGCCGGATCTCCTGCTATTAAGAAAGGGATATTACAAACTGTTAAAGTTGTGGATGAATTAGTGAAAGTAATGGGAAGACATAAACCTGAAAACATTGTCATTGAGATGGCAAGAGAAAATCAAACTACACAAAAAGGACAGAAAAATAGTAGAGAACGTATGAAAAGAATAGAAGAGGGTATTAAAGAATTGGGTAGTCAAATATTGAAAGAACACCCAGTGGAAAATACCCAGTTGCAAAATGAAAAATTATATCTTTACTACCTTCAAAATGGACGTGATATGTATGTTGATCAGGAATTAGATATAAATAGACTTTCAGATTATGATGTAGATCATATAGTTCCACAATCTTTCTTGAAAGATGATTCCATAGACAATAAAGTATTAACTAGAAGTGATAAAAATAGAGGTAAAAGTGATAATGTCCCAAGTGAGGAAGTCGTCAAAAAGATGAAAAATTACTGGCGTCAACTTTTGAATGCTAAATTAATTACTCAAAGAAAATTTGATAATTTGACTAAAGCAGAAAGAGGTGGGCTTTCTGAATTAGATAAAGCCGGGTTCATTAAAAGACAATTGGTCGAAACTAGACAAATTACTAAACATGTTGCCCAAATTTTAGATTCCCGTATGAACACTAAGTATGACGAAAATGATAAGTTAATACGTGAGGTTAAAGTCATTACTTTAAAATCAAAACTTGTCTCTGATTTCAGAAAGGATTTCCAATTCTATAAAGTTAGAGAAATTAATAATTATCATCATGCTCATGATGCATATTTGAATGCTGTAGTTGGAACTGCTTTAATCAAGAAATACCCTAAATTAGAATCTGAATTTGTATATGGTGATTACAAAGTCTATGATGTTAGAAAGATGATTGCTAAATCAGAACAAGAAATTGGTAAAGCTACAGCTAAATACTTCTTTTACTCTAACATTATGAATTTCTTTAAAACAGAAATTACTTTGGCAAACGGTGAAATTAGAAAAAGACCTCTTATTGAAACAAATGGTGAGACTGGAGAGATAGTTTGGGACAAAGGGCGTGATTTCGCTACTGTTAGAAAAGTTTTATCAATGCCACAAGTTAACATTGTAAAGAAAACAGAGGTTCAAACTGGTGGTTTCTCAAAAGAAAGTATTTTGCCTAAAAGAAATAGTGATAAATTGATTGCCAGAAAAAAGGATTGGGATCCAAAGAAATATGGTGGTTTCGACTCACCAACCGTAGCCTATTCTGTTTTGGTTGTGGCAAAGGTTGAAAAGGGTAAAAGTAAAAAGCTTAAATCAGTAAAAGAACTTTTGGGTATTACAATAATGGAAAGAAGTTCCTTTGAAAAGAACCCTATTGATTTTTTGGAAGCTAAAGGTTATAAGGAAGTAAAGAAGGACTTAATAATCAAATTGCCTAAATATTCTTTATTTGAATTAGAAAATGGGAGAAAAAGAATGTTGGCTTCTGCTGGAGAATTGCAAAAGGGTAATGAATTAGCATTGCCTTCCAAATATGTTAACTTCTTGTATTTAGCTTCACACTATGAAAAGTTGAAAGGGTCACCAGAAGATAACGAGCAAAAACAATTATTTGTTGAACAACACAAACACTACTTAGATGAGATTATAGAACAAATTAGTGAATTCAGTAAAAGAGTGATATTAGCTGATGCAAATTTAGATAAAGTTTTGTCAGCCTATAACAAACATAGAGATAAGCCAATTAGAGAACAAGCAGAAAACATTATTCACTTATTTACCCTTACCAATTTAGGAGCACCTGCTGCTTTCAAGTATTTTGATACAACAATTGATCGTAAAAGATATACCTCAACAAAAGAAGTCTTAGACGCCACCTTAATTCATCAATCAATCACTGGATTGTATGAGACAAGAATTGATTTGTCTCAATTGGGTGGTGAT
Protein Sequence
MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKNLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESFLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKFRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQIGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLRKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGNSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYFTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDSVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERLKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRHKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYLYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPSEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHVAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLNAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTEITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGITIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNELALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLDATLIHQSITGLYETRIDLSQLGGD
SV40 NLS — nuclear localization signal of SV40 (si… 10874..10895 Fwd
DNA Sequence
CCTAAGAAGAAAAGAAAAGTT
Protein Sequence
PKKKRKV
SV40 NLS — nuclear localization signal of SV40 (si… 10898..10919 Fwd
DNA Sequence
CCAAAGAAAAAGCGTAAGGTG
Protein Sequence
PKKKRKV
SV40 NLS — nuclear localization signal of SV40 (si… 10922..10943 Fwd
DNA Sequence
CCTAAGAAAAAGAGAAAGGTT
Protein Sequence
PKKKRKV
3xFLAG — three tandem FLAG(R) epitope tags, foll… 10943..11009 Fwd
DNA Sequence
GACTACAAAGACCATGACGGTGATTATAAAGATCATGACATCGACTACAAGGATGACGATGACAAG
Protein Sequence
DYKDHDGDYKDHDIDYKDDDDK
promoter 1
Label Gene Product Position Strand
AmpR promoter bla — 3551..3643 Rev
DNA Sequence
TTTGTTTATTTTTCTAAATACATTCAAATATGTATCCGCTCATGAGACAATAACCCTGATAAATGCTTCAATAATATTGAAAAAGGAAGAAT
protein_bind 3
Label Gene Product Position Strand
loxP — — 1088..1122 Fwd
DNA Sequence
ATAACTTCGTATAATGTATGCTATACGAAGTTAT
FRT (minimal) — — 1191..1225 Rev
DNA Sequence
GAAGTTCCTATTCTCTAGAAAGTATAGGAACTTC
FRT (minimal) — — 3935..3969 Fwd
DNA Sequence
GAAGTTCCTATTCTCTAGAAAGTATAGGAACTTC
rep_origin 1
Label Gene Product Position Strand
ori — — 1980..2569 Rev
DNA Sequence
TTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGCAAACAAAAAAACCACCGCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTTTTTCCGAAGGTAACTGGCTTCAGCAGAGCGCAGATACCAAATACTGTTCTTCTAGTGTAGCCGTAGTTAGCCCACCACTTCAAGAACTCTGTAGCACCGCCTACATACCTCGCTCTGCTAATCCTGTTACCAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGACGATAGTTACCGGATAAGGCGCAGCGGTCGGGCTGAACGGGGGGTTCGTGCACACAGCCCAGCTTGGAGCGAACGACCTACACCGAACTGAGATACCTACAGCGTGAGCTATGAGAAAGCGCCACGCTTCCCGAAGGGAGAAAGGCGGACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGGAGAGCGCACGAGGGAGCTTCCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTTCGCCACCTCTGACTTGAGCGTCGATTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATGGAAA
terminator 2
Label Gene Product Position Strand
TEF terminator — — 828..1026 Fwd
DNA Sequence
ACTGACAATAAAAAGATTCTTGTTTTCAAGAACTTGTCATTTGTATAGTTTTTTTATATTGTAGTTGTTCTATTTTAATCAAATGTTAGCGTGATTTATATTTTTTTTCGCCTCGACATCATCTGCCCAGATGCGAAGTTAAGTGCGCAGAAAGTAATATCATGCGTTAATCGTATGTGAATGCTGGTCGCTATACTG
CYC1 terminator S. cerevisiae CYC1 — 11119..11308 Fwd
DNA Sequence
ATCCGCTCTAACCGAAAAGGAAGGAGTTAGACAACCTGAAGTCTAGGTCCCTATTTATTTTTTTATAGTTATGTTAGTATTAAGAACGTTATTTATATTTCAAATTTTTCTTTTTTTTCTGTACAGACGCGTGTACGCATGTAACATTATACTGAAAACCTTGCTTGAGAAGGTTTTGGGACGCTCGAA