LOCUS       sequence-194008-        5577 bp    DNA     circular SYN 14-APR-2025
DEFINITION  To generate three copies of the HA epitope tag on cDNAs..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5577)
  TITLE     Wright lab backbones
REFERENCE   2  (bases 1 to 5577)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: pcDNA3-neo-cterminal-3HA
FEATURES             Location/Qualifiers
     source          1..5577
                     /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"
     CDS             995..1084
                     /codon_start=1
                     /product="three tandem HA epitope tags"
                     /label=3xHA
                     /translation="YPYDVPDYAGYPYDVPDYAGSYPYDVPDYA"
     promoter        complement(1129..1147)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1130..1147)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1167..1184)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1173..1397
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1443..1871
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1530..1549)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1740..1761
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(1880..1900)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        1885..2215
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2066..2201
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2128..2147
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2282..3076
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2336..2355)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2946..2965
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3250..3371
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3287..3306)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3341..3360
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3420..3436)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3420..3436)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3433..3455)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3444..3460
                     /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(3468..3474,3475..3492,3493..3498))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3513..3534
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3651..3668)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3822..4410)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3902..3921)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4581..5372,5373..5441))
                     /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     5204..5223
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5442..5546)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtcgactct 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 atgggtggac 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 gcttggtacc
      901 gagctcggat ccactagtaa cggccgccag tgtgctggaa ttctgcagat atccatcaca
      961 ctggcggccg ctcgaggatc tgggccgcat cttttaccca tacgatgttc ctgactatgc
     1021 gggctatccc tatgacgtcc cggactatgc aggatcctat ccatatgacg ttccagatta
     1081 cgctgcttag tgaggcccag atccaaataa taatctagag ggccctattc tatagtgtca
     1141 cctaaatgct agagctcgct gatcagcctc gactgtgcct tctagttgcc agccatctgt
     1201 tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc
     1261 ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg
     1321 tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc atgctgggga
     1381 tgcggtgggc tctatggctt ctgaggcgga aagaaccagc tggggctcta gggggtatcc
     1441 ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac
     1501 cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc
     1561 cacgttcgcc ggctttcccc gtcaagctct aaatcggggc atccctttag ggttccgatt
     1621 tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg
     1681 gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag
     1741 tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt
     1801 ataagggatt ttggggattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt
     1861 taacgcgaat taattctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc
     1921 ccaggcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca ggtgtggaaa
     1981 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
     2041 catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc
     2101 tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg cctctgcctc
     2161 tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaaaagct
     2221 cccgggagct tgtatatcca ttttcggatc tgatcaagag acaggatgag gatcgtttcg
     2281 catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg agaggctatt
     2341 cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt tccggctgtc
     2401 agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc tgaatgaact
     2461 gcaggacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt gcgcagctgt
     2521 gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag tgccggggca
     2581 ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg ctgatgcaat
     2641 gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag cgaaacatcg
     2701 catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg atctggacga
     2761 agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcgc gcatgcccga
     2821 cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca tggtggaaaa
     2881 tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc gctatcagga
     2941 catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg ctgaccgctt
     3001 cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct atcgccttct
     3061 tgacgagttc ttctgagcgg gactctgggg ttcgaaatga ccgaccaagc gacgcccaac
     3121 ctgccatcac gagatttcga ttccaccgcc gccttctatg aaaggttggg cttcggaatc
     3181 gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct ggagttcttc
     3241 gcccacccca acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca
     3301 aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc
     3361 aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc gtaatcatgg
     3421 tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc
     3481 ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg
     3541 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc
     3601 ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact
     3661 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
     3721 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
     3781 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
     3841 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
     3901 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
     3961 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcaatgc
     4021 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
     4081 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
     4141 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
     4201 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
     4261 aggacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
     4321 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
     4381 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
     4441 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
     4501 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
     4561 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
     4621 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
     4681 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
     4741 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     4801 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     4861 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
     4921 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     4981 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     5041 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     5101 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     5161 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
     5221 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     5281 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     5341 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     5401 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     5461 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     5521 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtc
//
