LOCUS       sequence-349355-        4442 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Donor vector for endogenous tagging of human GRN at the C-terminus 
            with halotag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4442)
  TITLE     iNDI donor vectors for endogenous tagging with Halotag
REFERENCE   2  (bases 1 to 4442)
  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"
COMMENT     Sequence Label: GRN_Halo_C_allele
FEATURES             Location/Qualifiers
     source          1..4442
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    5..26
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(143..160)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(314..902)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(394..413)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1073..1864,1865..1933))
                     /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     1696..1715
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1934..2038)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2106..2124
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(2162..2184)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2284..2303
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2497..2519
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2511..2528
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2512..2528
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             3009..3896
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="AEIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSSY
                     VWRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVIH
                     DWGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLIID
                     QNVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALVE
                     EYMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDLI
                     GSEIARWLSTLEISG"
     primer_bind     complement(4354..4370)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4354..4370)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4367..4389)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4378..4394
                     /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)."
     promoter        complement(join(4402..4408,4409..4426,4427..4432))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
ORIGIN
        1 cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg
       61 gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
      121 gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
      181 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
      241 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
      301 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
      361 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
      421 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
      481 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
      541 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
      601 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
      661 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
      721 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
      781 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
      841 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
      901 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
      961 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     1021 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     1081 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     1141 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     1201 caatgatacc gcgagatcca cgctcaccgg ctccagattt atcagcaata aaccagccag
     1261 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     1321 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     1381 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     1441 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     1501 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     1561 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     1621 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     1681 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     1741 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     1801 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     1861 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     1921 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     1981 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     2041 catttccccg aaaagtgcca cctgacgtct aagaaaccat tattatcatg acattaacct
     2101 ataaaaatag gcgtatcacg aggccctttc gtctcgcgcg tttcggtgat gacggtgaaa
     2161 acctctgaca catgcagctc ccggagacgg tcacagcttg tctgtaagcg gatgccggga
     2221 gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc tggcttaact
     2281 atgcggcatc agagcagatt gtactgagag tgcaccatat gcggtgtgaa ataccgcaca
     2341 gatgcgtaag gagaaaatac cgcatcaggc gccattcgcc attcaggctg cgcaactgtt
     2401 gggaagggcg atcggtgcgg gcctcttcgc tattacgcca gctggcgaaa gggggatgtg
     2461 ctgcaaggcg attaagttgg gtaacgccag ggttttccca gtcacgacgt tgtaaaacga
     2521 cggccagtga attcgagctc ggtacctcgc gaatgcatct agatgaaggc tcgatcctgc
     2581 gagaaggaag tggtctctgc ccagcctgcc accttcctgg cccgtagccc tcacgtgggt
     2641 gtgaaggacg tggagtgtgg ggaaggacac ttctgccatg ataaccagac ctgctgccga
     2701 gacaaccgac agggctgggc ctgctgtccc taccgccagg tcagtgccaa cccccatcct
     2761 ggggctgggt atggccaggg accaggtccc acctcgtcca accctctcgc ccccctctga
     2821 ccatccaggg cgtctgttgt gctgatcggc gccactgctg tcctgctggc ttccgctgcg
     2881 cagccagggg taccaagtgt ttgcgcaggg aggccccgcg ctgggacgcc cctttgaggg
     2941 acccagcctt gagacagctg ctgggtggag gaggttctgg aggcggtgga agtggtggcg
     3001 gaggtagcgc cgagatcggc accggctttc ccttcgatcc tcactacgtg gaagtgctgg
     3061 gcgagagaat gcactatgtg gacgtgggcc ccagagatgg aacccctgtg ctgtttctgc
     3121 acggcaaccc taccagcagc tacgtgtggc ggaacatcat ccctcacgtg gcccctacac
     3181 acagatgtat cgcccctgac ctgatcggca tgggcaagag cgataagccc gacctgggct
     3241 acttcttcga cgaccacgtg cggttcatgg acgcctttat tgaggccctc ggcctggaag
     3301 aggtggtgct ggttattcac gattggggca gcgccctggg attccactgg gccaagagaa
     3361 accccgagag agtgaaggga atcgccttca tggaattcat caggcccatt cctacctggg
     3421 acgagtggcc tgagttcgcc agagagacat tccaggcctt cagaaccacc gacgtgggca
     3481 gaaagctgat catcgaccag aacgtgttca tcgagggcac cctgcctatg ggagtcgtca
     3541 gaccactgac cgaggtggaa atggaccact acagagagcc ctttctgaac cccgtggacc
     3601 gggaacctct ttggagattc cccaacgagc tgcccattgc cggcgagcct gctaatattg
     3661 tggccctggt ggaagagtac atggactggc tgcatcagag ccccgtgcct aagctgctgt
     3721 tttggggaac acccggcgtg ctgattcctc ctgctgaagc tgccagactg gccaagagcc
     3781 tgcctaattg caaggccgtg gatatcggcc ctggcctgaa tctgctgcaa gaggacaacc
     3841 ccgatctgat cggctccgag attgccagat ggctgagcac cctggaaatc agcggataag
     3901 ggacagtact gaagactctg cagccctcgg gaccccactc ggagggtgcc ctctgctcag
     3961 gcctccctag cacctccccc taaccaaatt ctccctggac cccattctga gctccccatc
     4021 accatgggag gtggggcctc aatctaaggc cttccctgtc agaagggggt tgtggcaaaa
     4081 gccacattac aagctgccat cccctccccg tttcagtgga ccctgtggcc aggtgctttt
     4141 ccctatccac aggggtgttt gtgtgtgtgc gcgtgtgcgt ttcaataaag tttgtacact
     4201 ttcttaacag tgtctgattt gccgccctgc ctgccctccc cagggcccca gaacaggggt
     4261 tcacgtccac tgccaacact cccctcccct accccacagg gatcccgggc ccgtcgactg
     4321 cagaggcctg catgcaagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg
     4381 ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg
     4441 tg
//
