LOCUS       sequence-322681-        5046 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses a ST6GAL1 mutant with 21 amino acids in its TMD in 
            mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5046)
  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 5046)
  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: TMD21
FEATURES             Location/Qualifiers
     source          1..5046
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        70..375
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        376..579
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     530..550
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      996..1005
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1002..1004,1005..1007,1008..1721)
                     /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(1047..1068)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     polyA_signal    1843..1964
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1880..1899)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1934..1953
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1971..2426)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2108..2129)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2320..2339
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2453..2557
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2559..2916
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2559..2750
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2582..2602)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2767..2902
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2951..3745
                     /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             2951..3745
                     /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(3005..3024)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3615..3634
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3933..3952)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3977..4024
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4353..4941
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4842..4861
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gccatgcatt agttattaat agtaatcaat tacggggtca ttagttcata gcccatatat
       61 ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc
      121 ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca
      181 ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta
      241 tcatatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat
      301 tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc
      361 atcgctatta ccatggtgat gcggttttgg cagtacatca atgggcgtgg atagcggttt
      421 gactcacggg gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac
      481 caaaatcaac gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc
      541 ggtaggcgtg tacggtggga ggtctatata agcagagctg gtttagtgaa ccgtcagatc
      601 cgctagcgct accggactca gatctcgagg ccaccatgat tcacaccaac ctgaagaaaa
      661 agttcagctg ctgcgtacta gtcgcagtac tggtctttct tctgtttgca gtgatatgtg
      721 tgtggaagga aaagaagaaa gggagttact atgattcctt taaattgcaa accaaggaat
      781 tccaggtgtt aaagagtctg gggaaattgg ccatggggtc tgattcccag tctgtatcct
      841 caagcagcac ccaggacccc cacaggggcc gccagaccct cggcagtctc agaggcctag
      901 ccaaggccaa accagaggcc tccttccagg tgtggaacaa ggacagctct tccaaaaacc
      961 ttatccctag gctgcaaaag atctgggcac cggtcgccac catggtgtcc aagggcgagg
     1021 agctgttcac cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca
     1081 agttctccgt gcggggcgag ggcgagggcg atgccaccaa cggcaagctg accctgaagt
     1141 tcatcagcac caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgacct
     1201 acggcgtgca gagcttctcc cgctaccccg accacatgaa gcgccacgac ttcttcaaga
     1261 gcgccatgcc cgaaggctac gtccaggagc gcaccatctc cttcaaggac gacggcacct
     1321 acaagacccg cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga
     1381 agggcatcga cttcaaggag gacggcaaca tcctggggca caagctggag tacaacttca
     1441 actcccacaa cgtctatatc accgccgaca agcagaagaa cggcatcaag gccaacttca
     1501 agatccgcca caacgtggag gacggctccg tgcagctcgc cgaccactac cagcagaaca
     1561 cccccatcgg cgacggcccc gtgctgctgc ccgacaacca ctacctgtcc acccagtcca
     1621 agctgtccaa agaccccaac gagaagcgcg atcacatggt ccttctggaa ttcgtgaccg
     1681 ccgccgggat cactcacggc atggacgagc tgtacaagta agcggccgcg actctagatc
     1741 ataatcagcc ataccacatt tgtagaggtt ttacttgctt taaaaaacct cccacacctc
     1801 cccctgaacc tgaaacataa aatgaatgca attgttgttg ttaacttgtt tattgcagct
     1861 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca
     1921 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttaaggcgt aaattgtaag
     1981 cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca
     2041 ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga tagggttgag
     2101 tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca acgtcaaagg
     2161 gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct aatcaagttt
     2221 tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc cccgatttag
     2281 agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc
     2341 gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc
     2401 gcttaatgcg ccgctacagg gcgcgtcagg tggcactttt cggggaaatg tgcgcggaac
     2461 ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc
     2521 ctgataaatg cttcaataat attgaaaaag gaagagtcct gaggcggaaa gaaccagctg
     2581 tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     2641 caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca
     2701 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact
     2761 ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta
     2821 atttttttta tttatgcaga ggccgaggcc gcctcggcct ctgagctatt ccagaagtag
     2881 tgaggaggct tttttggagg cctaggcttt tgcaaagatc gatcaagaga caggatgagg
     2941 atcgtttcgc atgattgaac aagatggatt gcacgcaggt tctccggccg cttgggtgga
     3001 gaggctattc ggctatgact gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt
     3061 ccggctgtca gcgcaggggc gcccggttct ttttgtcaag accgacctgt ccggtgccct
     3121 gaatgaactg caagacgagg cagcgcggct atcgtggctg gccacgacgg gcgttccttg
     3181 cgcagctgtg ctcgacgttg tcactgaagc gggaagggac tggctgctat tgggcgaagt
     3241 gccggggcag gatctcctgt catctcacct tgctcctgcc gagaaagtat ccatcatggc
     3301 tgatgcaatg cggcggctgc atacgcttga tccggctacc tgcccattcg accaccaagc
     3361 gaaacatcgc atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg atcaggatga
     3421 tctggacgaa gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgag
     3481 catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat
     3541 ggtggaaaat ggccgctttt ctggattcat cgactgtggc cggctgggtg tggcggaccg
     3601 ctatcaggac atagcgttgg ctacccgtga tattgctgaa gagcttggcg gcgaatgggc
     3661 tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca tcgccttcta
     3721 tcgccttctt gacgagttct tctgagcggg actctggggt tcgaaatgac cgaccaagcg
     3781 acgcccaacc tgccatcacg agatttcgat tccaccgccg ccttctatga aaggttgggc
     3841 ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg
     3901 gagttcttcg cccaccctag ggggaggcta actgaaacac ggaaggagac aataccggaa
     3961 ggaacccgcg ctatgacggc aataaaaaga cagaataaaa cgcacggtgt tgggtcgttt
     4021 gttcataaac gcggggttcg gtcccagggc tggcactctg tcgatacccc accgagaccc
     4081 cattggggcc aatacgcccg cgtttcttcc ttttccccac cccacccccc aagttcgggt
     4141 gaaggcccag ggctcgcagc caacgtcggg gcggcaggcc ctgccatagc ctcaggttac
     4201 tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag
     4261 atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg
     4321 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
     4381 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
     4441 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt
     4501 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
     4561 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
     4621 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
     4681 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
     4741 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
     4801 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
     4861 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
     4921 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
     4981 tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt
     5041 attacc
//
