LOCUS       sequence-408101-        7709 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CcdB Dropout vector for part 1 of Hemagglutinin surface expression 
            in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7709)
  TITLE     Human expression plasmids for Hemagglutinin surface expression
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7709)
  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"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: HA Display H3_A/Hong Kong/1/1968 Part 1-DO
FEATURES             Location/Qualifiers
     source          1..7709
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    1004..1125
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1041..1060)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1095..1114
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(1174..1190)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1174..1190)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1187..1209)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1198..1214
                     /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(1222..1228,1229..1246,1247..1252))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1267..1288
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1405..1422)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1576..2164)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1656..1675)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2335..3126,3127..3195))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTHLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     2958..2977
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3196..3299)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(3375..3394)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        3566..3945
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3946..4149
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4100..4120
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    4151..4169
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4172..4190
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     4200..4224
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          4343..4475
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     4458..4477
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        4520..4538
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     sig_peptide     4584..4643
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
                     /translation="METDTLLLWVLLLWVPGSTG"
     CDS             complement(5007..5312)
                     /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="MQFKVYTYKRESRYRLFVDVQSDIIDTPG*RMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     complement(5109..5128)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             6631..6696
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             6703..6726
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             6733..6756
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             6799..6945
                     /codon_start=1
                     /product="transmembrane domain from platelet derived growth
                     factor receptor beta"
                     /label=PDGFR-beta TM domain
                     /translation="AVGQDTQEVIVVPHSLPFKVVVISAILALVVLTIISLIILIMLWQ
                     KKPR"
     primer_bind     complement(6992..7009)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6998..7222
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(7355..7374)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    7506..7553
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             join(7561..7709,1..874)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             join(7561..7709,1..874)
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
ORIGIN
        1 tagctgcgcc gatggtttct acaaagatcg ttatgtttat cggcactttg catcggccgc
       61 gctcccgatt ccggaagtgc ttgacattgg ggaattcagc gagagcctga cctattgcat
      121 ctcccgccgt gcacagggtg tcacgttgca agacctgcct gaaaccgaac tgcccgctgt
      181 tctgcagccg gtcgcggagg ccatggatgc gatcgctgcg gccgatctta gccagacgag
      241 cgggttcggc ccattcggac cgcaaggaat cggtcaatac actacatggc gtgatttcat
      301 atgcgcgatt gctgatcccc atgtgtatca ctggcaaact gtgatggacg acaccgtcag
      361 tgcgtccgtc gcgcaggctc tcgatgagct gatgctttgg gccgaggact gccccgaagt
      421 ccggcacctc gtgcacgcgg atttcggctc caacaatgtc ctgacggaca atggccgcat
      481 aacagcggtc attgactgga gcgaggcgat gttcggggat tcccaatacg aggtcgccaa
      541 catcttcttc tggaggccgt ggttggcttg tatggagcag cagacgcgct acttcgagcg
      601 gaggcatccg gagcttgcag gatcgccgcg gctccgggcg tatatgctcc gcattggtct
      661 tgaccaactc tatcagagct tggttgacgg caatttcgat gatgcagctt gggcgcaggg
      721 tcgatgcgac gcaatcgtcc gatccggagc cgggactgtc gggcgtacac aaatcgcccg
      781 cagaagcgcg gccgtctgga ccgatggctg tgtagaagta ctcgccgata gtggaaaccg
      841 acgccccagc actcgtccga gggcaaagga atagcacgta ctacgagatt tcgattccac
      901 cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat
      961 cctccagcgc ggggatctca tgctggagtt cttcgcccac cccaacttgt ttattgcagc
     1021 ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc
     1081 actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg tctgtatacc
     1141 gtcgacctct agctagagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg
     1201 ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg
     1261 tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc
     1321 gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt
     1381 gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct
     1441 gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga
     1501 taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
     1561 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
     1621 ctcaagtcgg aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
     1681 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
     1741 tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt
     1801 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
     1861 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
     1921 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
     1981 cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct
     2041 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
     2101 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
     2161 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     2221 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta
     2281 aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca
     2341 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
     2401 ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc
     2461 tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc
     2521 agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat
     2581 taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt
     2641 tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
     2701 cggttcccaa cgatcaaggt gagttacatg atcccccatg ttgtgcaaaa aagcggttag
     2761 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
     2821 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
     2881 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
     2941 cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
     3001 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
     3061 gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
     3121 tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa
     3181 atgttgaata ctcatactct tcctttttca atattattga agcatttatc agggttattg
     3241 tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgca
     3301 cacatttccc cgaaaagtgc cacctgacgt cgacggatcg ggagatctcc cgatccccta
     3361 tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagta tctgctccct
     3421 gcttgtgtgt tggaggtcgc tgagtagtgc gcgagcaaaa tttaagctac aacaaggcaa
     3481 ggcttgaccg acaattgcat gaagaatctg cttagggtta ggcgttttgc gctgcttcgc
     3541 gatgtacggg ccagatatac gcgttgacat tgattattga ctagttatta atagtaatca
     3601 attacggggt cattagttca tagcccatat atggagttcc gcgttacata acttacggta
     3661 aatggcccgc ctggctgacc gcccaacgac ccccgcccat tgacgtcaat aatgacgtat
     3721 gttcccatag taacgccaat agggactttc cattgacgtc aatgggtgga gtatttacgg
     3781 taaactgccc acttggcagt acatcaagtg tatcatatgc caagtacgcc ccctattgac
     3841 gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt acatgacctt atgggacttt
     3901 cctacttggc agtacatcta cgtattagtc atcgctatta ccatggtgat gcggttttgg
     3961 cagtacatca atgggcgtgg atagcggttt gactcacggg gatttccaag tctccacccc
     4021 attgacgtca atgggagttt gttttggcac caaaatcaac gggactttcc aaaatgtcgt
     4081 aacaactccg ccccattgac gcaaatgggc ggtaggcgtg tacggtggga ggtctatata
     4141 agcagagctc tccctatcag tgatagagat ctccctatca gtgatagaga tcgtcgacga
     4201 gctcgtttag tgaaccgtca gatcgcctgg agacgccatc cacgctgttt tgacctccat
     4261 agaagacacc gggaccgatc cagcctccgg actctagcgt ttaaacttaa gctgcagaag
     4321 ttggtcgtga ggcactgggc aggtaagtat caaggttaca agacaggttt aaggagacca
     4381 atagaaactg ggcttgtcga gacagagaag actcttgcgt ttctgatagg cacctattgg
     4441 tcttactgac atccactttg cctttctctc cacaggtgtc cactcccagt tcaattacag
     4501 ctcttaaggc tagagtactt aatacgactc actataggct agcctcgaga attcacgcgt
     4561 ggtacctcta gagtcgaccc gggatggaga cagacacact cctgctatgg gtactgctgc
     4621 tctgggttcc aggttccact ggtgacggtg gcaggtggag tcgaccgaca gccttccaaa
     4681 tgttcttctc aaacggaatc gtcgtatcca gcctactcgc tattgtcctc aatgccgtat
     4741 taaatcataa aaagaaataa gaaaaagagg tgcgagcctc ttttttgtgt gacaaaataa
     4801 aaacatctac ctattcatat acgctagtgt catagtcctg aaaatcatct gcatcaagaa
     4861 caatttcaca actcttatac ttttctctta caagtcgttc ggcttcatct ggattttcag
     4921 cctctatact tactaaacgt gataaagttt ctgtaatttc tactgtatcg acctgcagac
     4981 tggctgtgta taagggagcc tgacatttat attccccaga acatcaggtt aatggcgttt
     5041 ttgatgtcat tttcgcggtg gctgagatca gccacttctt ccccgataac ggagaccggc
     5101 acactggcca tatcggtggt catcatgcgc cagctttcat ccccgatatg caccaccggg
     5161 taaagttcac gggagacttt atctgacagc agacgtgcac tggccagggg gatcaccatc
     5221 cgtcacccgg gcgtgtcaat aatatcactc tgtacatcca caaacagacg ataacggctc
     5281 tctcttttat aggtgtaaac cttaaactgc atttcaccag cccctgttct cgtcagcaaa
     5341 agagccgttc atttcaataa accgggcgac ctcagccatc ccttcctgat tttccgcttt
     5401 ccagcgttcg gcacgcagac gacgggcttc attctgcatg gttgtgctta ccagaccgga
     5461 gatattgaca tcatatatgc cttgagcaac tgatagctgt cgctgtcaac tgtcactgta
     5521 atacgctgct tcatagcaca cctctttttg acacctgcca cccaccaccc cagcaccaac
     5581 caggagcaga ccagcctgta cgtgcaggcc agcggccggg tgaccgtgag cacccggcgg
     5641 agccagcaga ccatcatccc caacatcggc agccggccct gggtgcgggg cctgagcagc
     5701 cggatcagca tctactggac catcgtgaag cccggcgacg tgctggtgat caacagcaac
     5761 ggcaacctga tcgccccccg gggctacttc aagatgcgga ccggcaagag cagcatcatg
     5821 cggagcgacg cccccatcga tacatgtatc agcgagtgca tcacccccaa cggcagcatc
     5881 cccaacgaca agcccttcca gaacgtgaac aagatcacct acggcgcctg ccccaagtac
     5941 gtgaagcaga acaccctgaa gctggccacc ggcatgcgga acgtgcccga gaagcagacc
     6001 cggggcctgt tcggcgccat cgccggcttc atcgagaacg gctgggaggg catgatcgac
     6061 ggctggtacg gcttccggca ccagaacagc gagggcaccg gccaggccgc cgacctgaag
     6121 agcacccagg ccggcatcga ccagatcaac ggcaagctga accgggtgat cgagaagacc
     6181 aacgagaagt tccaccagat cgagaaggag ttcagcgagg tggagggccg gatccaggac
     6241 ctggagaagt acgtggagga caccaagatc gacctgtgga gctacaacgc cgagctgctg
     6301 gtggccctgg agaaccagca caccatcgac ctgaccgaca gcgagatgaa caagctgttc
     6361 gagaagaccc ggcggcagct gcgggagaac gccgaggaca tgggcaacgg ctgcttcaag
     6421 atctaccaca agtgcgacaa cgcctgcatc gagagcatcc ggaacggcac ctacgaccac
     6481 gacgtgtacc gggacgaggc cctgaacaac cggttccaga tcaagggcgt gcagggatca
     6541 ggatacatcc ccgaggcccc cagagatggc caggcctacg tgcggaagga cggcgagtgg
     6601 gtactgctga gcacattcct gggcagatcc gactacaaag accatgacgg tgattataaa
     6661 gatcatgaca tcgattacaa ggatgacgat gacaagagcg cctggtccca cccccagttc
     6721 gagaagggat ccctggaggt gctgttccag ggcccaggcg gcggcggtag tggagggggc
     6781 ggatctggcg gctcagctgc tgtgggccag gacacgcagg aggtcatcgt ggtgccacac
     6841 tccttgccct ttaaggtggt ggtgatctca gccatcctgg ccctggtggt gctcaccatc
     6901 atctccctta tcatcctcat catgctttgg cagaagaagc cacgttaacg gccgctcgag
     6961 tctagagggc ccgtttaaac ccgctgatca gcctcgactg tgccttctag ttgccagcca
     7021 tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc
     7081 ctttcctaat aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg
     7141 gggggtgggg tggggcagga cagcaagggg gaggattggg aagacaatag caggcatgct
     7201 ggggatgcgg tgggctctat ggcttctgag gcggaaagaa ccagctgggg ctctaggggg
     7261 tatccccacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     7321 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt
     7381 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
     7441 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt
     7501 acctagaagt tcctattccg aagttcctat tctctagaaa gtataggaac ttccttggcc
     7561 aaaaagcctg aactcaccgc gacgtctgtc gagaagtttc tgatcgaaaa gttcgacagc
     7621 gtctccgacc tgatgcagct ctcggagggc gaagaatctc gtgctttcag cttcgatgta
     7681 ggagggcgtg gatatgtcct gcgggtaaa
//
