LOCUS       sequence-197675-        5430 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Homology arms and linker-mEGFP sequence for C-terminus tagging of 
            human ST6GAL1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5430)
  AUTHORS   Allen Institute for Cell Science
  TITLE     Allen Institute for Cell Science Plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5430)
  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: 
            "Unpublished"
COMMENT     Sequence Label: AICSDP-26:ST6GAL1-mEGFP
FEATURES             Location/Qualifiers
     source          1..5430
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     134..153
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     393..414
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1522..1539)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1522..1538)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1531..1553)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(1747..1766)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1866..1888
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1926..1944)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2012..2116
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2117..2185,2186..2977)
                     /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     complement(2335..2354)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3148..3736
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3637..3656
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3890..3907
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4024..4045
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4060..4065,4066..4083,4084..4090)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4098..4114
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     4103..4125
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4122..4138
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4122..4138
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             complement(join(5170..5430,1..453,454..456))
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(5215..5236)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
ORIGIN
        1 gatatagacg ttgtggctgt tgtagttgta ctccagcttg tgccccagga tgttgccgtc
       61 ctccttgaag tcgatgccct tcagctcgat gcggttcacc agggtgtcgc cctcgaactt
      121 cacctcggcg cgggtcttgt agttgccgtc gtccttgaag aagatggtgc gctcctggac
      181 gtagccttcg ggcatggcgg acttgaagaa gtcgtgctgc ttcatgtggt cggggtagcg
      241 gctgaagcac tgcacgccgt aggtcagggt ggtcacgagg gtgggccagg gcacgggcag
      301 cttgccggtg gtgcagatga acttcagggt cagcttgccg taggtggcat cgccctcgcc
      361 ctcgccggac acgctgaact tgtggccgtt tacgtcgccg tccagctcga ccaggatggg
      421 caccaccccg gtgaacagct cctcgccctt gctcacggtg gccacaggtg ggtcccgagc
      481 ccgtggcacg gtgctctgga ggcagtgaat tgtgcgaaag ccaggcagtg tggcttttcc
      541 aagcaggtag atgtcctcat ctgtgccctg gttgagatgc ttcaccaaat tcttctcata
      601 gagcagcggg tggtaggcac ccatcgtgca ggcactatcg aagaacttct ggtagtagta
      661 gcacacgtca gtcttgcgct tggatgggag gaactcataa atatccacct ggtcacacag
      721 cgtcatcatg atgatgatac ctggagaggg gagcagaggt gagtcatgcc tgacaaccca
      781 ccccagcacc ccagagctct gcccagtggg ttcagcagct catgacccca aatttggaga
      841 gctttccctc tggagtttct atttatgact gtgattccca atctaagttt cagcaagcca
      901 agcccagcag tgttccttag gatgcagggc tgtgtgaggg ttatacacct ttgaatcccc
      961 agcacctagg ccagggcctg gtgaataata gatcctccct aaatgctagt agattaaatg
     1021 actccatcaa tcagccaatg aatgtatcca tggaataatt tatttggaga aagaggacaa
     1081 gattactgag ctgattgagg agacagtggc atcatgtgta gttagcactt tctagcactg
     1141 tgacatcctg cagaacctct tggatggtgc tatttgggag cagatttttc cttatagcat
     1201 ttgcatgaaa cagtgttggg gagggaagac ttttcttccc ttagctgggc aaagggacag
     1261 ctggaacagg agcactccac agtctgaatg atgagatgta gagaaaatcc aagaaggggg
     1321 cccctggcca gcattgttcc aataaactct gcacagcact gttactgatc ctatctttat
     1381 ttaaatactt gatatttcac tcatcataat ttttacatta attgtgtttt ctgaagatac
     1441 tgtagtgaaa tattatttat cttgatgact gagtttttaa ggggtacgcc cttaaattgt
     1501 aatcccaata cgcgtcaatt cactggccgt cgttttacaa cgtcgtgact gggaaaaccc
     1561 tggcgttacc caacttaatc gccttgcagc acatccccct ttcgccagct ggcgtaatag
     1621 cgaagaggcc cgcaccgatc gcccttccca acagttgcgc agcctgaatg gcgaatggcg
     1681 cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca tatggtgcac
     1741 tctcagtaca atctgctctg atgccgcata gttaagccag ccccgacacc cgccaacacc
     1801 cgctgacgcg ccctgacggg cttgtctgct cccggcatcc gcttacagac aagctgtgac
     1861 cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac gcgcgagacg
     1921 aaagggcctc gtgatacgcc tatttttata ggttaatgtc atgataataa tggtttctta
     1981 gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta
     2041 aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata
     2101 ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc
     2161 ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga
     2221 agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct
     2281 tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg
     2341 tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta
     2401 ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat
     2461 gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt
     2521 acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga
     2581 tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga
     2641 gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga
     2701 actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc
     2761 aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc
     2821 cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg
     2881 tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat
     2941 cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata
     3001 tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct
     3061 ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga
     3121 ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg
     3181 cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc
     3241 aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct
     3301 agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc
     3361 tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt
     3421 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg
     3481 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct
     3541 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag
     3601 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag
     3661 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg
     3721 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg
     3781 gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac
     3841 cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt
     3901 gagcgaggaa gcggaagagc gcccaatacg caaaccgcct ctccccgcgc gttggccgat
     3961 tcattaatgc agctggcacg acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc
     4021 aattaatgtg agttagctca ctcattaggc accccaggct ttacacttta tgcttccggc
     4081 tcgtatgttg tgtggaattg tgagcggata acaatttcac acaggaaaca gctatgacca
     4141 tgattacgcc aagctcggcg cgccattggg atgtaggagg tttcctaaaa aaatatcttg
     4201 agtctctaag ttcaggtaga tcttagaaaa taaacatggg aagaaagaaa aacacaattc
     4261 aaacggctca gatgtaatct gcttgccatg tttcttctct tcctctaaga ctcatctgaa
     4321 tgccccctgt gctataaagc cttctctgat ccacaaacac agttttaagt gcgtggccct
     4381 cctgggtgca cagagagagc aagcccatct ctctccaaat ccccagtgca ctgagcacac
     4441 tgaatcctac ctgcctgttc agatgtctgc ctctctcact gaaccgtagc aacttgagga
     4501 tagacactgt atccttctct accttccatc tccacatgga gagggaaagt gggctaacca
     4561 agacccttta atcataagca cccagtctaa ggaccagccc aaccagccag agtgatagga
     4621 acacgttcca ggggcatgga cctcgggagc aacttttctc tggaggcgcg gagcatgttt
     4681 ccaatcccct gtaccaccta cttgcccccc agagccaccc tgcattgggc acaattgtaa
     4741 tttaagagaa tttctttaaa acgtgaaatc aatatttaga tgcagacttc aagcaatata
     4801 gagcattggt ttgtgagaat ggcatcattc tgctgggaaa aacaaagccc agctttggca
     4861 gccctgatgg gagagataga ccttgcatct accaaggtgg aaggcgggga ggatgctgtc
     4921 tggaaagaat gctagatatg tgtgcatgtg tgtggcagga ggctctgggc tcataaactg
     4981 tctggctaca gggaaggctg gacctgattt cctgaccaca ggctcttgaa gtccccatgg
     5041 tcttgctgac agaggcccct agagtaaaag gagcaggctg gaattgttca ggaaaatgtt
     5101 cttcccaggc tggggtggcc aagagaccat tcatttaatg cctgatggag aagagtgagg
     5161 agggtgtgct tacttgtaca gctcgtccat gccgagagtg atcccggcgg cggtcacgaa
     5221 ctccagcagg accatgtgat cgcgcttctc gttggggtct ttgctcagct tggactgggt
     5281 gctcaggtag tggttgtcgg gcagcagcac ggggccgtcg ccgatggggg tgttctgctg
     5341 gtagtggtcg gcgagctgca cgctgccgtc ctcgatgttg tggcggatct tgaagttcac
     5401 cttgatgccg ttcttctgct tgtcggccat
//
