LOCUS       sequence-440021-        7366 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Gateway compatible empty vector (transient mammalian expression) 
            with a C-terminal 3xHA tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7366)
  TITLE     Harper Lab plasmids
REFERENCE   2  (bases 1 to 7366)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: pcDNA3.1-ccdB-3xHA
FEATURES             Location/Qualifiers
     source          1..7366
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    969..1093
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(1118..1123,1124..1141,1142..1148)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             1202..1861
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(1268..1287)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             2203..2508
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     2387..2406
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(2549..2673)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             2675..2755
                     /codon_start=1
                     /product="three tandem HA epitope tags"
                     /label=3xHA
                     /translation="YPYDVPDYAYPYDVPDYAYPYDVPDYA"
     primer_bind     complement(2798..2815)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2804..2915
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3064..3492
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3151..3170)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3361..3382
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3501..3521)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3506..3835
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3686..3821
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3748..3767
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3884..4909
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             3884..4909
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     polyA_signal    5039..5160
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5076..5095)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5130..5149
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5209..5225)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5209..5225)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5222..5244)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5233..5249
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(5257..5263,5264..5281,5282..5287))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5302..5323
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5440..5457)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5611..6199)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5691..5710)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6370..7161,7162..7230))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6993..7012
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7231..7335)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaactt aagcttggta ccgagctcgg atccactagt ccagtgtggt ggaattctgc
      961 agatatcaac aagtttgtac aaaaaagctg aacgagaaac gtaaaatgat ataaatatca
     1021 atatattaaa ttagattttg cataaaaaac agactacata atactgtaaa acacaacata
     1081 tccagtcact atggcggccg cattaggcac cccaggcttt acactttatg cttccggctc
     1141 gtataatgtg tggattttga gttaggatcc gtcgagattt tcaggagcta aggaagctaa
     1201 aatggagaaa aaaatcactg gatataccac cgttgatata tcccaatggc atcgtaaaga
     1261 acattttgag gcatttcagt cagttgctca atgtacctat aaccagaccg ttcagctgga
     1321 tattacggcc tttttaaaga ccgtaaagaa aaataagcac aagttttatc cggcctttat
     1381 tcacattctt gcccgcctga tgaatgctca tccggaattc cgtatggcaa tgaaagacgg
     1441 tgagctggtg atatgggata gtgttcaccc ttgttacacc gttttccatg agcaaactga
     1501 aacgttttca tcgctctgga gtgaatacca cgacgatttc cggcagtttc tacacatata
     1561 ttcgcaagat gtggcgtgtt acggtgaaaa cctggcctat ttccctaaag ggtttattga
     1621 gaatatgttt ttcgtctcag ccaatccctg ggtgagtttc accagttttg atttaaacgt
     1681 ggccaatatg gacaacttct tcgcccccgt tttcaccatg ggcaaatatt atacgcaagg
     1741 cgacaaggtg ctgatgccgc tggcgattca ggttcatcat gccgtttgtg atggcttcca
     1801 tgtcggcaga atgcttaatg aattacaaca gtactgcgat gagtggcagg gcggggcgta
     1861 aagatctgga tccggcttac taaaagccag ataacagtat gcgtatttgc gcgctgattt
     1921 ttgcggtata agaatatata ctgatatgta tacccgaagt atgtcaaaaa gaggtatgct
     1981 atgaagcagc gtattacagt gacagttgac agcgacagct atcagttgct caaggcatat
     2041 atgatgtcaa tatctccggt ctggtaagca caaccatgca gaatgaagcc cgtcgtctgc
     2101 gtgccgaacg ctggaaagcg gaaaatcagg aagggatggc tgaggtcgcc cggtttattg
     2161 aaatgaacgg ctcttttgct gacgagaaca ggggctggtg aaatgcagtt taaggtttac
     2221 acctataaaa gagagagccg ttatcgtctg tttgtggatg tacagagtga tattattgac
     2281 acgcccgggc gacggatggt gatccccctg gccagtgcac gtctgctgtc agataaagtc
     2341 tcccgtgaac tttacccggt ggtgcatatc ggggatgaaa gctggcgcat gatgaccacc
     2401 gatatggcca gtgtgccggt ctccgttatc ggggaagaag tggctgatct cagccaccgc
     2461 gaaaatgaca tcaaaaacgc cattaacctg atgttctggg gaatataaat gtcaggctcc
     2521 cttatacaca gccagtctgc aggtcgacca tagtgactgg atatgttgtg ttttacagta
     2581 ttatgtagtc tgttttttat gcaaaatcta atttaatata ttgatattta tatcatttta
     2641 cgtttctcgt tcagctttct tgtacaaagt ggtttaccca tacgatgttc cagattacgc
     2701 ttatccttat gacgtacctg actatgcata cccttatgat gtaccagact acgcttgaaa
     2761 ctcgagtcta gagggcccgt ttaaacccgc tgatcagcct cgactgtgcc ttctagttgc
     2821 cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc
     2881 actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct
     2941 attctggggg gtggggtggg gcaggacagc aaggggaaga caatagcagg catgctgggg
     3001 atgcggtggg ctctatggct tctgaggcgg aaagaaccag ctggggctct agggggtatc
     3061 cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
     3121 ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg
     3181 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
     3241 ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg
     3301 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
     3361 gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt
     3421 tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat
     3481 ttaacgcgaa ttaattctgt ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc
     3541 cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca ggtgtggaaa
     3601 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
     3661 catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc
     3721 tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg cctctgcctc
     3781 tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaaaagct
     3841 cccgggagct tgtatatcca ttttcggatc tgatcagcac gtgatgaaaa agcctgaact
     3901 caccgcgacg tctgtcgaga agtttctgat cgaaaagttc gacagcgtct ccgacctgat
     3961 gcagctctcg gagggcgaag aatctcgtgc tttcagcttc gatgtaggag ggcgtggata
     4021 tgtcctgcgg gtaaatagct gcgccgatgg tttctacaaa gatcgttatg tttatcggca
     4081 ctttgcatcg gccgcgctcc cgattccgga agtgcttgac attggggaat tcagcgagag
     4141 cctgacctat tgcatctccc gccgtgcaca gggtgtcacg ttgcaagacc tgcctgaaac
     4201 cgaactgccc gctgttctgc agccggtcgc ggaggccatg gatgcgatcg ctgcggccga
     4261 tcttagccag acgagcgggt tcggcccatt cggaccgcaa ggaatcggtc aatacactac
     4321 atggcgtgat ttcatatgcg cgattgctga tccccatgtg tatcactggc aaactgtgat
     4381 ggacgacacc gtcagtgcgt ccgtcgcgca ggctctcgat gagctgatgc tttgggccga
     4441 ggactgcccc gaagtccggc acctcgtgca cgcggatttc ggctccaaca atgtcctgac
     4501 ggacaatggc cgcataacag cggtcattga ctggagcgag gcgatgttcg gggattccca
     4561 atacgaggtc gccaacatct tcttctggag gccgtggttg gcttgtatgg agcagcagac
     4621 gcgctacttc gagcggaggc atccggagct tgcaggatcg ccgcggctcc gggcgtatat
     4681 gctccgcatt ggtcttgacc aactctatca gagcttggtt gacggcaatt tcgatgatgc
     4741 agcttgggcg cagggtcgat gcgacgcaat cgtccgatcc ggagccggga ctgtcgggcg
     4801 tacacaaatc gcccgcagaa gcgcggccgt ctggaccgat ggctgtgtag aagtactcgc
     4861 cgatagtgga aaccgacgcc ccagcactcg tccgagggca aaggaatagc acgtgctacg
     4921 agatttcgat tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga
     4981 cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccccaa
     5041 cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa
     5101 taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta
     5161 tcatgtctgt ataccgtcga cctctagcta gagcttggcg taatcatggt catagctgtt
     5221 tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa
     5281 gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact
     5341 gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc
     5401 ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg
     5461 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
     5521 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag
     5581 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     5641 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     5701 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     5761 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
     5821 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     5881 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     5941 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     6001 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt
     6061 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     6121 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
     6181 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
     6241 gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta
     6301 gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg
     6361 gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg
     6421 ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc
     6481 atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc
     6541 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc
     6601 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
     6661 tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat
     6721 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
     6781 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
     6841 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
     6901 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
     6961 accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt
     7021 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
     7081 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
     7141 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
     7201 aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat
     7261 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca
     7321 aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtc
//
