LOCUS       sequence-348247-        5995 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Nhx1-iGFP  in yeast cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5995)
  AUTHORS   Casler JC, Johnson N, Krahn AH, Pantazopoulou A, Day KJ, Glick BS
  TITLE     Clathrin adaptors mediate two sequential pathways of intra-Golgi 
            recycling.
  JOURNAL   J Cell Biol. 2022 Jan 3;221(1). pii: 212747. doi: 
            10.1083/jcb.202103199. Epub 2021 Nov 5.
  PUBMED    34739034
REFERENCE   2  (bases 1 to 5995)
  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 
            Biol."; date: "2022-01-3"; volume: "221(1). pii: 212747. doi"; 
            pages: " 10.1083/jcb.202103199. Epub 2021 Nov 5"
COMMENT     Sequence Label: YIplac211-NHX1-iGFP
FEATURES             Location/Qualifiers
     source          1..5995
                     /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"
     primer_bind     complement(1076..1104)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     complement(2493..2510)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2493..2509)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2502..2524)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(2718..2737)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2837..2859
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2977..3780)
                     /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(3781..3996)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(4003..4021)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4089..4193
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4194..4262,4263..5054)
                     /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(4412..4431)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5225..5813
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5714..5733
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5967..5984
                     /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 ccaagcttgc
      241 atgcgcagct atttctatat gtgttgctcg ttggtgtgct gtgttcccat tgtcgcaatt
      301 tgttaactgg atatatagag taaagacaat cagatctatg agcggcataa ccggagaaaa
      361 tatttctgtt cccgatgaaa taccctacaa ttaccaaatg atgacatttt gggcaggttt
      421 acgtggtgct gttggtgtcg ccttggcgtt gggaattcaa ggtgagtata agttcacttt
      481 attggcaacg gtccttgttg ttgttgtttt aacagttatc atttttgggg gcactactgc
      541 aggaatgtta gaagttttaa atattaagac tggttgcata agtgaagaag atacatctga
      601 tgacgagttt gatatagagg ctccaagggc gataaattta ttgaacggta gttctattca
      661 gacagatttg ggcccatatt ctgacaacaa ttctccagat atttcaattg accaattcgc
      721 ggtcagcagt aacaagaatc tccccaataa catatccaca actggtggta atacttttgg
      781 aggccttaat gaaactgaga atactagccc taacccggca aggtcttcta tggataagcg
      841 taatttgaga gataaactgg gaacaatctt taattccgac tcacaatggt ttcaaaattt
      901 tgatgaacag gtattgaagc cagtattctt ggacaacgtt tctccatcct tacaagattc
      961 ggctacgcaa tcacctgcag atttctcttc ccaaaaccac acggatccag gaagttcggg
     1021 gtccagcggt tcttcaggca gttccatgga tagcactgag agccttttca ctggagttgt
     1081 cccaattctt gttgaattag atggtgatgt taatgggcac aaattttctg tcagtggaga
     1141 gggtgaaggt gatgcaacat acggaaaact tacccttaaa tttatttgca ctactggaaa
     1201 actacctgtt ccttggccaa cacttgtcac tactttaact tatggtgttc agtgcttttc
     1261 aagatacccg gatcatatga aacagcatga ctttttcaag agtgccatgc ccgaaggtta
     1321 tgtacaggaa agaactatat ttttcaaaga tgacgggaac tacaagacac gtgctgaagt
     1381 caagtttgaa ggtgataccc ttgttaatag aatcgagtta aaaggtattg attttaaaga
     1441 agatggaaac attcttgggc acaaattgga atacaactat aactcacaca atgtatacat
     1501 catggcagac aaacaaaaga atggaatcaa agctaacttc aaaattagac acaacattga
     1561 agatggaagc gttcaactag cagaccatta tcaacaaaat actccaattg gcgatggccc
     1621 tgtcctttta ccagacaacc attacctgtc cacacaatct gccctttcga aagatcccaa
     1681 cgaaaagaga gaccacatgg tccttaaaga gtttgtaaca gctgctggga ttacacatgg
     1741 cagctccggc tccagcgggt aaagcggccg cactttaaag tgtatggttt ccttgtttct
     1801 aatataaata tattaattaa gtctttatac attctctctt actcaacgcc gaaggaatac
     1861 ccttctacac tttattcatt ttaaatcatg tactgttata tacacgctag taggattgat
     1921 cttatttaat catatttcgc acgttacccg actgctttca gttttgccgc tttgtgataa
     1981 tgaggaattt cagctagctt ttaatgagcc gatactaaat caccatgatt ttgggagagg
     2041 aaaagaagaa aaggtaaaac aacgaatatt tttcaaatac cttagttact gaatatccat
     2101 aactactcgt ataattggtg agaaaatagt gcatatttag tttacttttt gcctttgatt
     2161 gaaaatatat attcatggtg ctttttactc ggtgtgaaaa ggcaagaaag gagaaactcg
     2221 ccgctggtta taagccctta gttgattatt tgattgattg tgatactccc acatttttag
     2281 aaagaattga ggcaatccaa gagtgggacc gatccagaga tgatctttat gtttggattc
     2341 ccattctaga cagaatggac gggctgttgt taaaagtggc agagaaatat aaatacaagc
     2401 aagaccccaa gaaggagtgt gaagtcaaat tggtggaaat ggaagcgcat gatgttgatt
     2461 attgtttgaa aatgggtacc gagctcgaat tcactggccg tcgttttaca acgtcgtgac
     2521 tgggaaaacc ctggcgttac ccaacttaat cgccttgcag cacatccccc tttcgccagc
     2581 tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg cagcctgaat
     2641 ggcgaatggc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc
     2701 atatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca gccccgacac
     2761 ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga
     2821 caagctgtga ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa
     2881 cgcgcgagac gaaagggcgg gtaataactg atataattaa attgaagctc taatttgtga
     2941 gtttagtata catgcattta cttataatac agttttttag ttttgctggc cgcatcttct
     3001 caaatatgct tcccagcctg cttttctgta acgttcaccc tctaccttag catcccttcc
     3061 ctttgcaaat agtcctcttc caacaataat aatgtcagat cctgtagaga ccacatcatc
     3121 cacggttcta tactgttgac ccaatgcgtc tcccttgtca tctaaaccca caccgggtgt
     3181 cataatcaac caatcgtaac cttcatctct tccacccatg tctctttgag caataaagcc
     3241 gataacaaaa tctttgtcgc tcttcgcaat gtcaacagta cccttagtat attctccagt
     3301 agatagggag cccttgcatg acaattctgc taacatcaaa aggcctctag gttcctttgt
     3361 tacttcttct gccgcctgct tcaaaccgct aacaatacct gggcccacca caccgtgtgc
     3421 attcgtaatg tctgcccatt ctgctattct gtatacaccc gcagagtact gcaatttgac
     3481 tgtattacca atgtcagcaa attttctgtc ttcgaagagt aaaaaattgt acttggcgga
     3541 taatgccttt agcggcttaa ctgtgccctc catggaaaaa tcagtcaaga tatccacatg
     3601 tgtttttagt aaacaaattt tgggacctaa tgcttcaact aactccagta attccttggt
     3661 ggtacgaaca tccaatgaag cacacaagtt tgtttgcttt tcgtgcatga tattaaatag
     3721 cttggcagca acaggactag gatgagtagc agcacgttcc ttatatgtag ctttcgacat
     3781 gatttatctt cgtttcctgc atgtttttgt tctgtgcagt tgggttaaga atactgggca
     3841 atttcatgtt tcttcaacac tacatatgcg tatatatacc aatctaagtc tgtgctcctt
     3901 ccttcgttct tccttctgtt cggagattac cgaatcaaaa aaatttcaag gaaaccgaaa
     3961 tcaaaaaaaa gaataaaaaa aaaatgatga attgaaaagc tggcctcgtg atacgcctat
     4021 ttttataggt taatgtcatg ataataatgg tttcttagac gtcaggtggc acttttcggg
     4081 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     4141 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta
     4201 ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg
     4261 ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg
     4321 gttacatcga actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac
     4381 gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg
     4441 acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt
     4501 actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg
     4561 ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac
     4621 cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt
     4681 gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag
     4741 caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc
     4801 aacaattaat agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc
     4861 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta
     4921 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg
     4981 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga
     5041 ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac
     5101 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
     5161 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     5221 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     5281 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     5341 gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc
     5401 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     5461 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     5521 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     5581 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     5641 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     5701 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     5761 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     5821 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc
     5881 ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg
     5941 ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaaga
//
