LOCUS       sequence-322682-        5049 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses a ST6GAL1 mutant with 22 amino acids in its TMD in 
            mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5049)
  AUTHORS   Sun X, Mahajan D, Chen B, Song Z, Lu L
  TITLE     A quantitative study of the Golgi retention of glycosyltransferases.
  JOURNAL   J Cell Sci. 2021 Oct 15;134(20). pii: 272560. doi: 
            10.1242/jcs.258564. Epub 2021 Oct 21.
  PUBMED    34533190
REFERENCE   2  (bases 1 to 5049)
  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 Cell 
            Sci."; date: "2021-10-15"; volume: "134(20). pii: 272560. doi"; 
            pages: " 10.1242/jcs.258564. Epub 2021 Oct 21"
COMMENT     Sequence Label: TMD22
FEATURES             Location/Qualifiers
     source          1..5049
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        97..402
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        403..606
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     557..577
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      1026..1035
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1032..1034,1035..1037,1038..1751)
                     /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(1077..1098)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     polyA_signal    1873..1994
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1910..1929)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1964..1983
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2001..2456)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2138..2159)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2350..2369
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2483..2587
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2589..2946
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2589..2780
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2612..2632)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2797..2932
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2981..3775
                     /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             2981..3775
                     /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(3035..3054)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3645..3664
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3963..3982)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4007..4054
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4383..4971
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4872..4891
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgattctg tggataaccg tattaccgcc atgcattagt tattaatagt aatcaattac
       61 ggggtcatta gttcatagcc catatatgga gttccgcgtt acataactta cggtaaatgg
      121 cccgcctggc tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc
      181 catagtaacg ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac
      241 tgcccacttg gcagtacatc aagtgtatca tatatgccaa gtacgccccc tattgacgtc
      301 aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct
      361 acttggcagt acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag
      421 tacatcaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt
      481 gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaac
      541 aactccgccc cattgacgca aatgggcggt aggcgtgtac ggtgggaggt ctatataagc
      601 agagctggtt tagtgaaccg tcagatccgc tagcgctacc ggactcagat ctcgaggcca
      661 ccatgattca caccaacctg aagaaaaagt tcagctgctg cgtactagtc gcagtactgg
      721 tcctatttct tctgtttgca gtgatatgtg tgtggaagga aaagaagaaa gggagttact
      781 atgattcctt taaattgcaa accaaggaat tccaggtgtt aaagagtctg gggaaattgg
      841 ccatggggtc tgattcccag tctgtatcct caagcagcac ccaggacccc cacaggggcc
      901 gccagaccct cggcagtctc agaggcctag ccaaggccaa accagaggcc tccttccagg
      961 tgtggaacaa ggacagctct tccaaaaacc ttatccctag gctgcaaaag atctgggcac
     1021 cggtcgccac catggtgtcc aagggcgagg agctgttcac cggggtggtg cccatcctgg
     1081 tcgagctgga cggcgacgta aacggccaca agttctccgt gcggggcgag ggcgagggcg
     1141 atgccaccaa cggcaagctg accctgaagt tcatcagcac caccggcaag ctgcccgtgc
     1201 cctggcccac cctcgtgacc accctgacct acggcgtgca gagcttctcc cgctaccccg
     1261 accacatgaa gcgccacgac ttcttcaaga gcgccatgcc cgaaggctac gtccaggagc
     1321 gcaccatctc cttcaaggac gacggcacct acaagacccg cgccgaggtg aagttcgagg
     1381 gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca
     1441 tcctggggca caagctggag tacaacttca actcccacaa cgtctatatc accgccgaca
     1501 agcagaagaa cggcatcaag gccaacttca agatccgcca caacgtggag gacggctccg
     1561 tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc
     1621 ccgacaacca ctacctgtcc acccagtcca agctgtccaa agaccccaac gagaagcgcg
     1681 atcacatggt ccttctggaa ttcgtgaccg ccgccgggat cactcacggc atggacgagc
     1741 tgtacaagta agcggccgcg actctagatc ataatcagcc ataccacatt tgtagaggtt
     1801 ttacttgctt taaaaaacct cccacacctc cccctgaacc tgaaacataa aatgaatgca
     1861 attgttgttg ttaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc
     1921 acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc
     1981 atcaatgtat cttaaggcgt aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa
     2041 tttttgttaa atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa
     2101 atcaaaagaa tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact
     2161 attaaagaac gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc
     2221 actacgtgaa ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa
     2281 tcggaaccct aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc
     2341 gagaaaggaa gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt
     2401 cacgctgcgc gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtcagg
     2461 tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc
     2521 aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag
     2581 gaagagtcct gaggcggaaa gaaccagctg tggaatgtgt gtcagttagg gtgtggaaag
     2641 tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc
     2701 aggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     2761 tagtcagcaa ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt
     2821 tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc
     2881 gcctcggcct ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt
     2941 tgcaaagatc gatcaagaga caggatgagg atcgtttcgc atgattgaac aagatggatt
     3001 gcacgcaggt tctccggccg cttgggtgga gaggctattc ggctatgact gggcacaaca
     3061 gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct
     3121 ttttgtcaag accgacctgt ccggtgccct gaatgaactg caagacgagg cagcgcggct
     3181 atcgtggctg gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc
     3241 gggaagggac tggctgctat tgggcgaagt gccggggcag gatctcctgt catctcacct
     3301 tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg cggcggctgc atacgcttga
     3361 tccggctacc tgcccattcg accaccaagc gaaacatcgc atcgagcgag cacgtactcg
     3421 gatggaagcc ggtcttgtcg atcaggatga tctggacgaa gagcatcagg ggctcgcgcc
     3481 agccgaactg ttcgccaggc tcaaggcgag catgcccgac ggcgaggatc tcgtcgtgac
     3541 ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt ctggattcat
     3601 cgactgtggc cggctgggtg tggcggaccg ctatcaggac atagcgttgg ctacccgtga
     3661 tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc
     3721 cgctcccgat tcgcagcgca tcgccttcta tcgccttctt gacgagttct tctgagcggg
     3781 actctggggt tcgaaatgac cgaccaagcg acgcccaacc tgccatcacg agatttcgat
     3841 tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg
     3901 atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccctag ggggaggcta
     3961 actgaaacac ggaaggagac aataccggaa ggaacccgcg ctatgacggc aataaaaaga
     4021 cagaataaaa cgcacggtgt tgggtcgttt gttcataaac gcggggttcg gtcccagggc
     4081 tggcactctg tcgatacccc accgagaccc cattggggcc aatacgcccg cgtttcttcc
     4141 ttttccccac cccacccccc aagttcgggt gaaggcccag ggctcgcagc caacgtcggg
     4201 gcggcaggcc ctgccatagc ctcaggttac tcatatatac tttagattga tttaaaactt
     4261 catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc
     4321 ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct
     4381 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta
     4441 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc
     4501 ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac
     4561 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct
     4621 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat
     4681 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg
     4741 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
     4801 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg
     4861 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga
     4921 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc
     4981 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct
     5041 gcgttatcc
//
