LOCUS       sequence-442501-        5528 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expression of mGM-CSF in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5528)
  AUTHORS   Heller L, Pottinger C, Jaroszeski MJ, Gilbert R, Heller R
  TITLE     In vivo electroporation of plasmids encoding GM-CSF or interleukin-2
            into existing B16 melanomas combined with electrochemotherapy 
            induces long-term antitumour immunity.
  JOURNAL   Melanoma Res. 2000 Dec;10(6):577-83.
  PUBMED    11198480
REFERENCE   2  (bases 1 to 5528)
  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: "Melanoma 
            Res."; date: "2000-12"; volume: "10(6)"; pages: "577-83"
COMMENT     Sequence Label: pCR3.1-mGM-CSF
FEATURES             Location/Qualifiers
     source          1..5528
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        5..384
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        385..588
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     539..559
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     633..652
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        633..651
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1270..1287)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1276..1500
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      complement(1632..2220)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1712..1731)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    2549..2596
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     2621..2640
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     CDS             complement(2828..3622)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             complement(2828..3622)
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2939..2958)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     3549..3568
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(3657..4014)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3671..3806
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        3823..4014
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     3971..3991
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     CDS             complement(join(4074..4865,4866..4934))
                     /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     4697..4716
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4935..5039)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      5066..5521
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5153..5172)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5363..5384
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 cgttgacatt gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat
       61 agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg
      121 cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata
      181 gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta
      241 catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc
      301 gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca gtacatctac
      361 gtattagtca tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga
      421 tagcggtttg actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg
      481 ttttggcacc aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg
      541 caaatgggcg gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact
      601 agagaaccca ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa
      661 gctggctagc gtttaaactt aagcttggta ccgagctcgg atccactagt ccagtgtggt
      721 ggaattcggc ttctgaggag gatgtggctg cagaatttac ttttcctggg cattgtggtc
      781 tacagcctct cagcacccac ccgctcaccc atcactgtca cccggccttg gaagcatgta
      841 gaggccatca aagaagccct gaacctcctg gatgacatgc ctgtcacgtt gaatgaagag
      901 gtagaagtcg tctctaacga gttctccttc aagaagctaa catgtgtgca gacccgcctg
      961 aagatattcg agcagggtct acggggcaat ttcaccaaac tcaagggcgc cttgaacatg
     1021 acagccagct actaccagac atactgcccc ccaactccgg aaacggactg tgaaacacaa
     1081 gttaccacct atgcggattt catagacagc cttaaaacct ttctgactga tatccccttt
     1141 gaatgcaaaa aaccaggcca aaaatgagga agcccaggcc agctctgaat ccagaagccg
     1201 aattctgcag atatccagca cagtggcggc cgctcgagtc tagagggccc gtttaaaccc
     1261 gctgatcagc ctcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg
     1321 tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa
     1381 ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca
     1441 gcaaggggga ggattgggaa gacaatagca ggcatgctgg ggatgcggtg ggctctatgg
     1501 cttctgaggc ggaaagaacc agtggcggta atacggttat ccacagaatc aggggataac
     1561 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     1621 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     1681 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     1741 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     1801 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     1861 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     1921 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     1981 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     2041 aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg
     2101 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     2161 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
     2221 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     2281 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
     2341 tgaagtttta aatcaatcta aagtatatat gagtaacctg aggctatggc agggcctgcc
     2401 gccccgacgt tggctgcgag ccctgggcct tcacccgaac ttggggggtg gggtggggaa
     2461 aaggaagaaa cgcgggcgta ttggccccaa tggggtctcg gtggggtatc gacagagtgc
     2521 cagccctggg accgaacccc gcgtttatga acaaacgacc caacaccgtg cgttttattc
     2581 tgtcttttta ttgccgtcat agcgcgggtt ccttccggta ttgtctcctt ccgtgtttca
     2641 gttagcctcc ccctagggtg ggcgaagaac tccagcatga gatccccgcg ctggaggatc
     2701 atccagccgg cgtcccggaa aacgattccg aagcccaacc tttcatagaa ggcggcggtg
     2761 gaatcgaaat ctcgtgatgg caggttgggc gtcgcttggt cggtcatttc gaaccccaga
     2821 gtcccgctca gaagaactcg tcaagaaggc gatagaaggc gatgcgctgc gaatcgggag
     2881 cggcgatacc gtaaagcacg aggaagcggt cagcccattc gccgccaagc tcttcagcaa
     2941 tatcacgggt agccaacgct atgtcctgat agcggtccgc cacacccagc cggccacagt
     3001 cgatgaatcc agaaaagcgg ccattttcca ccatgatatt cggcaagcag gcatcgccat
     3061 gggtcacgac gagatcctcg ccgtcgggca tgctcgcctt gagcctggcg aacagttcgg
     3121 ctggcgcgag cccctgatgc tcttcgtcca gatcatcctg atcgacaaga ccggcttcca
     3181 tccgagtacg tgctcgctcg atgcgatgtt tcgcttggtg gtcgaatggg caggtagccg
     3241 gatcaagcgt atgcagccgc cgcattgcat cagccatgat ggatactttc tcggcaggag
     3301 caaggtgaga tgacaggaga tcctgccccg gcacttcgcc caatagcagc cagtcccttc
     3361 ccgcttcagt gacaacgtcg agcacagctg cgcaaggaac gcccgtcgtg gccagccacg
     3421 atagccgcgc tgcctcgtct tgcagttcat tcagggcacc ggacaggtcg gtcttgacaa
     3481 aaagaaccgg gcgcccctgc gctgacagcc ggaacacggc ggcatcagag cagccgattg
     3541 tctgttgtgc ccagtcatag ccgaatagcc tctccaccca agcggccgga gaacctgcgt
     3601 gcaatccatc ttgttcaatc atgcgaaacg atcctcatcc tgtctcttga tcgatctttg
     3661 caaaagccta ggcctccaaa aaagcctcct cactacttct ggaatagctc agaggccgag
     3721 gcggcctcgg cctctgcata aataaaaaaa attagtcagc catggggcgg agaatgggcg
     3781 gaactgggcg gagttagggg cgggatgggc ggagttaggg gcgggactat ggttgctgac
     3841 taattgagat gcatgctttg catacttctg cctgctgggg agcctgggga ctttccacac
     3901 ctggttgctg actaattgag atgcatgctt tgcatacttc tgcctgctgg ggagcctggg
     3961 gactttccac accctaactg acacacattc cacagctggt tctttccgcc tcaggactct
     4021 tcctttttca ataaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa
     4081 tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc
     4141 tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct
     4201 gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca
     4261 gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt
     4321 aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt
     4381 gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc
     4441 ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc
     4501 tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt
     4561 atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact
     4621 ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc
     4681 ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt
     4741 ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg
     4801 atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct
     4861 gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa
     4921 tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt
     4981 ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc
     5041 acatttcccc gaaaagtgcc acctgacgcg ccctgtagcg gcgcattaag cgcggcgggt
     5101 gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc
     5161 gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg
     5221 gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat
     5281 tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg
     5341 ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct
     5401 atctcggtct attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa
     5461 aatgagctga tttaacaaaa atttaacgcg aattttaaca aaatattaac gcttacaatt
     5521 tacgcgcg
//
