LOCUS       sequence-492037-        6866 bp    DNA     circular SYN 06-NOV-2025
DEFINITION  Insert an N-terminal HaloTag at VRG4 by popin-popout.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6866)
  TITLE     Glick lab plasmids 2025
REFERENCE   2  (bases 1 to 6866)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Nov 6, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: YIplac211-HaloTag-Vrg4
FEATURES             Location/Qualifiers
     source          1..6866
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    106..127
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(142..147,148..165,166..172)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    180..196
                     /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     185..207
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     204..220
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     204..220
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             720..1610
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="MAEIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSS
                     YVWRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVI
                     HDWGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLII
                     DQNVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALV
                     EEYMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDL
                     IGSEIARWLSTLEISG"
     primer_bind     complement(3364..3381)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3364..3380)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3373..3395)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(3589..3608)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3708..3730
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3848..4651)
                     /codon_start=1
                     /gene="S. cerevisiae URA3"
                     /product="orotidine-5'-phosphate decarboxylase, required 
                     for uracil biosynthesis"
                     /label=URA3
                     /note="yeast auxotrophic marker, counterselectable with 
                     5-fluoroorotic acid (5-FOA)"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     promoter        complement(4652..4867)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(4874..4892)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4960..5064
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5065..5133,5134..5925)
                     /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(5283..5302)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6096..6684
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6585..6604
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6838..6855
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
       61 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg tgagttagct
      121 cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt tgtgtggaat
      181 tgtgagcgga taacaatttc acacaggaaa cagctatgac catgattacg ccaagcttgt
      241 aatggaccat ggcaatgtgc gccatatata tatgcggctc ccttatgagt agaagaaaat
      301 atatgactgg tggtacaaag tggcatggtg gtatatagat gtaggaacac catttgagtt
      361 tgccataatg attaaatttc cctttttata aagaattagc cgcccataac acgaaaaata
      421 tcacagttga cgaaagaaga cacgtcgcct aaacgcgtaa tgaatcgcaa tatacgaaaa
      481 gctaaacggg tttttcagaa tcctcttacc cagccttgga taatagtttt ggccgaaaaa
      541 aaaaaaaaga aagaaaaaaa ttgaaatttc gcgatccgaa caaacaatga acagaaaaac
      601 agaatatata taacgctgcc aagcctattt cttgttgccg attaattgtt gacatcctct
      661 taattaccaa aagagcctaa gaaaacaaac acactaacca cacagtatct ttcgcccgaa
      721 tggctgaaat cggtactggt ttcccttttg acccacacta tgtcgaagta ttaggtgaaa
      781 gaatgcacta tgttgatgtc ggtcctagag atggtactcc agttttgttt ttacatggta
      841 acccaacatc ttcatacgtt tggagaaaca tcatcccaca tgttgcacca actcatagat
      901 gtattgctcc agatttgatt ggtatgggta aatctgataa gccagatttg ggttatttct
      961 ttgatgatca tgttagattc atggatgctt ttattgaagc attgggtttg gaagaagttg
     1021 ttttggttat tcatgattgg ggttcagcat taggttttca ttgggctaag agaaacccag
     1081 aaagagttaa gggtatcgct tttatggaat tcattagacc aattccaact tgggatgaat
     1141 ggccagaatt cgcaagagaa acattccaag cttttagaac tacagatgtt ggtagaaagt
     1201 tgatcatcga tcaaaacgtt tttattgaag gtactttgcc aatgggtgtt gttagaccat
     1261 taacagaagt tgaaatggat cattacagag aaccattttt aaacccagtt gatagagaac
     1321 cattgtggag atttccaaat gaattaccaa ttgctggtga accagcaaac atcgttgctt
     1381 tggttgaaga atacatggat tggttacatc aatctccagt tccaaagttg ttattttggg
     1441 gtacaccagg tgttttgatt ccaccagcag aagctgcaag attggctaag tcattgccaa
     1501 actgtaaggc tgttgatatt ggtccaggtt taaatttgtt acaagaagat aatccagact
     1561 tgataggtag tgaaatagcc agatggttgt ccacattaga aatctcaggt ggttcttcag
     1621 aattgaaaac aggtcatgca ggccataacc cttgggcttc agttgccaat tccggtccga
     1681 tctctatttt atcctactgt ggttcctcta ttttaatgac ggtgactaac aagttcgtcg
     1741 tcaatttgaa ggatttcaac atgaactttg tcatgctttt cgtgcaatct ttggtttgta
     1801 ctataacctt gattatccta cgtatactgg gctatgcgaa gttccgttca ttaaacaaaa
     1861 cagacgccaa gaactggttc cctatttcct ttttactggt cttgatgatc tacacctctt
     1921 cgaaggcttt acaatacttg gctgttccaa tttacaccat tttcaagaat ttgactatta
     1981 tcttgattgc ttatggtgag gttctctttt ttggtggctc tgtcacctcc atggaattgt
     2041 catcattttt gttgatggtc ctttcttctg tcgttgcaac ttggggtgac cagcaagctg
     2101 tggctgccaa ggctgcttca ttggctgaag gagcagccgg tgctgttgcc tcctttaacc
     2161 caggttattt ctggatgttc accaactgta tcacttctgc attattcgtt cttataatga
     2221 gaaagagaat taagttaact aacttcaagg atttcgacac tatgttttac aacaatgttt
     2281 tggctctacc tattctattg ctgttttctt tctgtgtgga agattggtct tcagttaatt
     2341 tgaccaataa cttttctaac gattcgctaa ctgctatgat catcagtggt gttgcatccg
     2401 tcggtatttc ttactgttcc ggttggtgtg ttcgtgttac ttcgtctact acatattcga
     2461 tggtaggggc tttgaacaag ctgccaattg ccttgtctgg tttgattttc tttgatgctc
     2521 caagaaactt cttatctatt ctctccattt ttattggttt cctatcaggt attatttatg
     2581 ctgttgccaa acaaaagaag caacaagccc aacctttacg taaatgagaa cttacggggg
     2641 gtgcaattta tttttttttt ttggtttatt ttattttata gaggcatcta atgcaagtag
     2701 atttatatac aattatactt aaaattgata taccttagaa caggtcggac acaacgtgtg
     2761 gtaactccaa tatatcacta atgacaatag cgccttctgg tgtttgcccc aaaatttcat
     2821 ttacttcatt ctcaactaaa tgaatacatg ttttcatgcc caatttgata cctgtctcga
     2881 tgtttttccc actgtcatct ataaaataag cattctcgta tcttgccaac ccgctttcct
     2941 tcatggcttt ttcaaatgct ttaacatgtg gtttgcagac cagcgtatct gtcctagagt
     3001 agtcacaata tgtcaatcca tcaaataaat ccgctatacc caacaatctc aggcaacgta
     3061 ttgcgtggtt tttataagcg ttggtgaaaa gccacaattt atcaatcttt cccgattgtc
     3121 tcaatctcaa cagcatattc cttaatggta tatcaggttt caagatatct tgtagtggta
     3181 aggaatcatc cacaagccgg ttatattcca gtgcgttcac tttgtggaac attacaagtc
     3241 cccttatggc aagtccatac tctttgtagt aagagttatt caaaacgtga gcatcttcag
     3301 gtgatagttt caagtgtgtt tgaaagaacc tcagtataga ttgttgcata agatcgtgaa
     3361 ttcactggcc gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa
     3421 tcgccttgca gcacatcccc ctttcgccag ctggcgtaat agcgaagagg cccgcaccga
     3481 tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg cgcctgatgc ggtattttct
     3541 ccttacgcat ctgtgcggta tttcacaccg catatggtgc actctcagta caatctgctc
     3601 tgatgccgca tagttaagcc agccccgaca cccgccaaca cccgctgacg cgccctgacg
     3661 ggcttgtctg ctcccggcat ccgcttacag acaagctgtg accgtctccg ggagctgcat
     3721 gtgtcagagg ttttcaccgt catcaccgaa acgcgcgaga cgaaagggcg ggtaataact
     3781 gatataatta aattgaagct ctaatttgtg agtttagtat acatgcattt acttataata
     3841 cagtttttta gttttgctgg ccgcatcttc tcaaatatgc ttcccagcct gcttttctgt
     3901 aacgttcacc ctctacctta gcatcccttc cctttgcaaa tagtcctctt ccaacaataa
     3961 taatgtcaga tcctgtagag accacatcat ccacggttct atactgttga cccaatgcgt
     4021 ctcccttgtc atctaaaccc acaccgggtg tcataatcaa ccaatcgtaa ccttcatctc
     4081 ttccacccat gtctctttga gcaataaagc cgataacaaa atctttgtcg ctcttcgcaa
     4141 tgtcaacagt acccttagta tattctccag tagataggga gcccttgcat gacaattctg
     4201 ctaacatcaa aaggcctcta ggttcctttg ttacttcttc tgccgcctgc ttcaaaccgc
     4261 taacaatacc tgggcccacc acaccgtgtg cattcgtaat gtctgcccat tctgctattc
     4321 tgtatacacc cgcagagtac tgcaatttga ctgtattacc aatgtcagca aattttctgt
     4381 cttcgaagag taaaaaattg tacttggcgg ataatgcctt tagcggctta actgtgccct
     4441 ccatggaaaa atcagtcaag atatccacat gtgtttttag taaacaaatt ttgggaccta
     4501 atgcttcaac taactccagt aattccttgg tggtacgaac atccaatgaa gcacacaagt
     4561 ttgtttgctt ttcgtgcatg atattaaata gcttggcagc aacaggacta ggatgagtag
     4621 cagcacgttc cttatatgta gctttcgaca tgatttatct tcgtttcctg catgtttttg
     4681 ttctgtgcag ttgggttaag aatactgggc aatttcatgt ttcttcaaca ctacatatgc
     4741 gtatatatac caatctaagt ctgtgctcct tccttcgttc ttccttctgt tcggagatta
     4801 ccgaatcaaa aaaatttcaa ggaaaccgaa atcaaaaaaa agaataaaaa aaaaatgatg
     4861 aattgaaaag ctggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg
     4921 gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta
     4981 tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt
     5041 caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc
     5101 ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa
     5161 gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt
     5221 aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt
     5281 ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc
     5341 atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg
     5401 gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg
     5461 gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac
     5521 atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca
     5581 aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta
     5641 actggcgaac tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat
     5701 aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa
     5761 tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag
     5821 ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat
     5881 agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt
     5941 tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg
     6001 aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga
     6061 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     6121 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     6181 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     6241 gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     6301 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     6361 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     6421 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     6481 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     6541 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     6601 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     6661 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     6721 ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac
     6781 cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc
     6841 gagtcagtga gcgaggaagc ggaaga
//
