LOCUS       sequence-323646-        6086 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses a ST6GAL1 mutant with a GFP tag in its C-terminus in 
            mammalian cells. One wildtype Tac stem region is inserted within ST 
            and the other mutated Tac stem region is inserted between ST and 
            GFP..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6086)
  AUTHORS   Sun X, Tie HC, Chen B, Lu L
  TITLE     Glycans function as a Golgi export signal to promote the 
            constitutive exocytic trafficking.
  JOURNAL   J Biol Chem. 2020 Aug 21. pii: RA120.014476. doi: 
            10.1074/jbc.RA120.014476.
  PUBMED    32826314
REFERENCE   2  (bases 1 to 6086)
  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: "J Biol 
            Chem. 2020 Aug 21. pii: RA120.014476. doi: 
            10.1074/jbc.RA120.014476."
COMMENT     Sequence Label: ST-2xTstem(5A)-GFP
FEATURES             Location/Qualifiers
     source          1..6086
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      2026..2035
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2032..2034,2035..2037,2038..2751)
                     /codon_start=1
                     /product="monomeric GFP variant optimized for use in 
                     oxidizing environments such as the secretory pathway 
                     (Costantini et al., 2015)"
                     /label=moxGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL
                     KFISTTGKLPVPWPTLVTTLTYGVQSFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDD
                     GTYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIK
                     ANFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITHGMDELYK"
     primer_bind     complement(2077..2098)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     polyA_signal    2874..2995
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2911..2930)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2965..2984
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3002..3457)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3139..3160)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3351..3370
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3484..3588
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3590..3947
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3590..3781
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3613..3633)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3798..3933
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3982..4776
                     /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             3982..4776
                     /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(4036..4055)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4646..4665
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4964..4983)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5008..5055
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5384..5972
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5873..5892
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggactcag atctcgaggc caccatgatt cacaccaacc tgaagaaaaa gttcagctgc
      661 tgcgtcctgg tctttcttct gtttgcagtc atctgtgtgt ggaaggaaaa gaagaaaggg
      721 agttactatg attcctttaa attgcaaacc aaggaattcc aggtgttaaa gagtctgggg
      781 aaattggcca tggggtctga ttccccgcgg ctcgtcacaa caacagattt tcaaatacag
      841 acagaaatgg ctgcaaccat ggagacgtcc atatttacaa cagagtacca gggatcccag
      901 tctgtatcct caagcagcac ccaggacccc cacaggggcc gccagaccct cggcagtctc
      961 agaggcctag ccaaggccaa accagaggcc tccttccagg tgtggaacaa ggacagctct
     1021 tccaaaaacc ttatccctag gctgcaaaag atctggaaga attacctaag catgaacaag
     1081 tacaaagtgt cctacaaggg gccaggacca ggcatcaagt tcagtgcaga ggccctgcgc
     1141 tgccacctcc gggaccatgt gaatgtatcc atggtagagg tcacagattt tcccttcaat
     1201 acctctgaat gggagggtta tctgcccaag gagagcatta ggaccaaggc tgggccttgg
     1261 ggcaggtgtg ctgttgtgtc gtcagcggga tctctgaagt cctcccaact aggcagagaa
     1321 atcgatgatc atgacgcagt cctgaggttt aatggggcac ccacagccaa cttccaacaa
     1381 gatgtgggca caaaaactac cattcgcctg atgaactctc agttggttac cacagagaag
     1441 cgcttcctca aagacagttt gtacaatgaa ggaatcctaa ttgtatggga cccatctgta
     1501 taccactcag atatcccaaa gtggtaccag aatccggatt ataatttctt taacaactac
     1561 aagacttatc gtaagctgca ccccaatcag cccttttaca tcctcaagcc ccagatgcct
     1621 tgggagctat gggacattct tcaagaaatc tccccagaag agattcagcc aaacccccca
     1681 tcctctggga tgcttggtat catcatcatg atgacgctgt gtgaccaggt ggatatttat
     1741 gagttcctcc catccaagcg caagactgac gtgtgctact actaccagaa gttcttcgat
     1801 agtgcctgca cgatgggtgc ctaccacccg ctgctctatg agaagaattt ggtgaagcat
     1861 ctcaaccagg gcacagatga ggacatctac ctgcttggaa aagccacact gcctggcttc
     1921 cggaccattc actgcgcacc ggtcctcgtc acaacagcag attttcaaat acaggcagaa
     1981 atggctgcag ccatggagac gtccatattt gcagcagagt accaggccac catggtgtcc
     2041 aagggcgagg agctgttcac cggggtggtg cccatcctgg tcgagctgga cggcgacgta
     2101 aacggccaca agttctccgt gcggggcgag ggcgagggcg atgccaccaa cggcaagctg
     2161 accctgaagt tcatcagcac caccggcaag ctgcccgtgc cctggcccac cctcgtgacc
     2221 accctgacct acggcgtgca gagcttctcc cgctaccccg accacatgaa gcgccacgac
     2281 ttcttcaaga gcgccatgcc cgaaggctac gtccaggagc gcaccatctc cttcaaggac
     2341 gacggcacct acaagacccg cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc
     2401 atcgagctga agggcatcga cttcaaggag gacggcaaca tcctggggca caagctggag
     2461 tacaacttca actcccacaa cgtctatatc accgccgaca agcagaagaa cggcatcaag
     2521 gccaacttca agatccgcca caacgtggag gacggctccg tgcagctcgc cgaccactac
     2581 cagcagaaca cccccatcgg cgacggcccc gtgctgctgc ccgacaacca ctacctgtcc
     2641 acccagtcca agctgtccaa agaccccaac gagaagcgcg atcacatggt ccttctggaa
     2701 ttcgtgaccg ccgccgggat cactcacggc atggacgagc tgtacaagta aagcggccgc
     2761 gactctagat cataatcagc cataccacat ttgtagaggt tttacttgct ttaaaaaacc
     2821 tcccacacct ccccctgaac ctgaaacata aaatgaatgc aattgttgtt gttaacttgt
     2881 ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag
     2941 catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta tcttaaggcg
     3001 taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta aatcagctca
     3061 ttttttaacc aataggccga aatcggcaaa atcccttata aatcaaaaga atagaccgag
     3121 atagggttga gtgttgttcc agtttggaac aagagtccac tattaaagaa cgtggactcc
     3181 aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga accatcaccc
     3241 taatcaagtt ttttggggtc gaggtgccgt aaagcactaa atcggaaccc taaagggagc
     3301 ccccgattta gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa
     3361 gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc
     3421 acacccgccg cgcttaatgc gccgctacag ggcgcgtcag gtggcacttt tcggggaaat
     3481 gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg
     3541 agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtcc tgaggcggaa
     3601 agaaccagct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc tccccagcag
     3661 gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga aagtccccag
     3721 gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accatagtcc
     3781 cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc
     3841 atggctgact aatttttttt atttatgcag aggccgaggc cgcctcggcc tctgagctat
     3901 tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaagat cgatcaagag
     3961 acaggatgag gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc
     4021 gcttgggtgg agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat
     4081 gccgccgtgt tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg
     4141 tccggtgccc tgaatgaact gcaagacgag gcagcgcggc tatcgtggct ggccacgacg
     4201 ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta
     4261 ttgggcgaag tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta
     4321 tccatcatgg ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc
     4381 gaccaccaag cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc
     4441 gatcaggatg atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg
     4501 ctcaaggcga gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg
     4561 ccgaatatca tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt
     4621 gtggcggacc gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc
     4681 ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc
     4741 atcgccttct atcgccttct tgacgagttc ttctgagcgg gactctgggg ttcgaaatga
     4801 ccgaccaagc gacgcccaac ctgccatcac gagatttcga ttccaccgcc gccttctatg
     4861 aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg
     4921 atctcatgct ggagttcttc gcccacccta gggggaggct aactgaaaca cggaaggaga
     4981 caataccgga aggaacccgc gctatgacgg caataaaaag acagaataaa acgcacggtg
     5041 ttgggtcgtt tgttcataaa cgcggggttc ggtcccaggg ctggcactct gtcgataccc
     5101 caccgagacc ccattggggc caatacgccc gcgtttcttc cttttcccca ccccaccccc
     5161 caagttcggg tgaaggccca gggctcgcag ccaacgtcgg ggcggcaggc cctgccatag
     5221 cctcaggtta ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga
     5281 tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt
     5341 tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc
     5401 tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc
     5461 cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac
     5521 caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac
     5581 cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt
     5641 cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct
     5701 gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat
     5761 acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt
     5821 atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg
     5881 cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt
     5941 gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt
     6001 tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg
     6061 tggataaccg tattaccgcc atgcat
//
