LOCUS       sequence-365736-        5708 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  transient expression of C-terminal GFP tagged SLC35A1 isoform a in 
            mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5708)
  AUTHORS   Li D, Mukhopadhyay S
  TITLE     A three-pocket model for substrate coordination and selectivity by 
            the nucleotide sugar transporters SLC35A1 and SLC35A2.
  JOURNAL   J Biol Chem. 2021 Sep;297(3):101069. doi: 10.1016/j.jbc.2021.101069.
            Epub 2021 Aug 10.
  PUBMED    34384782
REFERENCE   2  (bases 1 to 5708)
  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."; date: "2021-09"; volume: "297(3):101069. doi"; pages: " 
            10.1016/j.jbc.2021.101069. Epub 2021 Aug 10"
COMMENT     Sequence Label: pEGFP.N3-SLC35A1-GFP
FEATURES             Location/Qualifiers
     source          1..5708
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        70..373
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        374..577
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     528..548
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      1657..1666
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1663..1665,1666..1668,1669..2382)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1663..1665,1666..1668,1669..2379)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1708..1729)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1969..1988)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2316..2337
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2505..2626
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2542..2561)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2596..2615
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2633..3088)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2770..2791)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2982..3001
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3115..3219
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3221..3578
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3221..3412
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3244..3264)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3429..3564
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3613..4407
                     /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             3613..4407
                     /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(3667..3686)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4277..4296
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4595..4614)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4639..4686
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5015..5603
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5504..5523
                     /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 tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta
      301 tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat
      361 cgctattacc atggtgatgc ggttttggca gtacatcaat gggcgtggat agcggtttga
      421 ctcacgggga tttccaagtc tccaccccat tgacgtcaat gggagtttgt tttggcacca
      481 aaatcaacgg gactttccaa aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg
      541 taggcgtgta cggtgggagg tctatataag cagagctggt ttagtgaacc gtcagatccg
      601 ctagcgctac cggactcaga tctcgagctc aagcttatgg ctgccccgag agacaatgtc
      661 actttattat tcaagttata ctgcttggca gtgatgaccc tgatggctgc agtctatacc
      721 atagctttaa gatacacaag gacatcagac aaagaactct acttttcaac cacagccgtg
      781 tgtatcacag aagttataaa gttattgcta agtgtgggaa ttttagctaa agaaactggt
      841 agtctgggta gattcaaagc atctttaaga gaaaatgtct tggggagccc caaggaactg
      901 ttgaagttaa gtgtgccatc gttagtgtat gctgttcaga acaacatggc tttcctagct
      961 cttagcaatc tggatgcagc agtgtaccag gtgacctacc agttgaagat tccgtgtact
     1021 gctttatgca ctgttttaat gttaaaccgg acactcagca aattacagtg ggtttcagtt
     1081 tttatgctgt gtgctggagt tacgcttgta cagtggaaac cagcccaagc tacaaaagtg
     1141 gtggtggaac aaaatccatt attagggttt ggcgctatag ctattgctgt attgtgctca
     1201 ggatttgcag gagtatattt tgaaaaagtt ttaaagagtt cagatacttc tctttgggtg
     1261 agaaacattc aaatgtatct atcagggatt attgtgacat tagctggcgt ctacttgtca
     1321 gatggagctg aaattaaaga aaaaggattt ttctatggtt acacatatta tgtctggttt
     1381 gtcatctttc ttgcaagtgt tggtggcctc tacacttctg ttgtggttaa gtacacagac
     1441 aacatcatga aaggcttttc tgcagcagcg gccattgtcc tttccaccat tgcttcagta
     1501 atgctgtttg gattacagat aacactcacc tttgccctgg gtactcttct tgtatgtgtt
     1561 tccatatatc tctatggatt acccagacaa gacactacat ccatccaaca aggagaaaca
     1621 gcttcaaagg agagagttat tggtgtggga tccatcgcca ccatggtgag caagggcgag
     1681 gagctgttca ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac
     1741 aagttcagcg tgtccggcga gggcgagggc gatgccacct acggcaagct gaccctgaag
     1801 ttcatctgca ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccctgacc
     1861 tacggcgtgc agtgcttcag ccgctacccc gaccacatga agcagcacga cttcttcaag
     1921 tccgccatgc ccgaaggcta cgtccaggag cgcaccatct tcttcaagga cgacggcaac
     1981 tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg
     2041 aagggcatcg acttcaagga ggacggcaac atcctggggc acaagctgga gtacaactac
     2101 aacagccaca acgtctatat catggccgac aagcagaaga acggcatcaa ggtgaacttc
     2161 aagatccgcc acaacatcga ggacggcagc gtgcagctcg ccgaccacta ccagcagaac
     2221 acccccatcg gcgacggccc cgtgctgctg cccgacaacc actacctgag cacccagtcc
     2281 gccctgagca aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc
     2341 gccgccggga tcactctcgg catggacgag ctgtacaagt aaagcggccg cgactctaga
     2401 tcataatcag ccataccaca tttgtagagg ttttacttgc tttaaaaaac ctcccacacc
     2461 tccccctgaa cctgaaacat aaaatgaatg caattgttgt tgttaacttg tttattgcag
     2521 cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt
     2581 cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttaaggc gtaaattgta
     2641 agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac
     2701 caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga gatagggttg
     2761 agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc caacgtcaaa
     2821 gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc ctaatcaagt
     2881 tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag cccccgattt
     2941 agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga
     3001 gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc
     3061 gcgcttaatg cgccgctaca gggcgcgtca ggtggcactt ttcggggaaa tgtgcgcgga
     3121 acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa
     3181 ccctgataaa tgcttcaata atattgaaaa aggaagagtc ctgaggcgga aagaaccagc
     3241 tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta
     3301 tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag
     3361 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa
     3421 ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc catggctgac
     3481 taattttttt tatttatgca gaggccgagg ccgcctcggc ctctgagcta ttccagaagt
     3541 agtgaggagg cttttttgga ggcctaggct tttgcaaaga tcgatcaaga gacaggatga
     3601 ggatcgtttc gcatgattga acaagatgga ttgcacgcag gttctccggc cgcttgggtg
     3661 gagaggctat tcggctatga ctgggcacaa cagacaatcg gctgctctga tgccgccgtg
     3721 ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca agaccgacct gtccggtgcc
     3781 ctgaatgaac tgcaagacga ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct
     3841 tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg actggctgct attgggcgaa
     3901 gtgccggggc aggatctcct gtcatctcac cttgctcctg ccgagaaagt atccatcatg
     3961 gctgatgcaa tgcggcggct gcatacgctt gatccggcta cctgcccatt cgaccaccaa
     4021 gcgaaacatc gcatcgagcg agcacgtact cggatggaag ccggtcttgt cgatcaggat
     4081 gatctggacg aagagcatca ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg
     4141 agcatgcccg acggcgagga tctcgtcgtg acccatggcg atgcctgctt gccgaatatc
     4201 atggtggaaa atggccgctt ttctggattc atcgactgtg gccggctggg tgtggcggac
     4261 cgctatcagg acatagcgtt ggctacccgt gatattgctg aagagcttgg cggcgaatgg
     4321 gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg catcgccttc
     4381 tatcgccttc ttgacgagtt cttctgagcg ggactctggg gttcgaaatg accgaccaag
     4441 cgacgcccaa cctgccatca cgagatttcg attccaccgc cgccttctat gaaaggttgg
     4501 gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc
     4561 tggagttctt cgcccaccct agggggaggc taactgaaac acggaaggag acaataccgg
     4621 aaggaacccg cgctatgacg gcaataaaaa gacagaataa aacgcacggt gttgggtcgt
     4681 ttgttcataa acgcggggtt cggtcccagg gctggcactc tgtcgatacc ccaccgagac
     4741 cccattgggg ccaatacgcc cgcgtttctt ccttttcccc accccacccc ccaagttcgg
     4801 gtgaaggccc agggctcgca gccaacgtcg gggcggcagg ccctgccata gcctcaggtt
     4861 actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga
     4921 agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag
     4981 cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa
     5041 tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag
     5101 agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg
     5161 ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat
     5221 acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta
     5281 ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg
     5341 gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc
     5401 gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa
     5461 gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc
     5521 tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt
     5581 caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct
     5641 tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc
     5701 gtattacc
//
