LOCUS       sequence-252101-        5660 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For labeling endoplasmic reticulum (ER) in S. cerevisiae with 
            GFP-HDEL.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5660)
  AUTHORS   Omer S, Greenberg SR, Lee WL
  TITLE     Cortical dynein pulling mechanism is regulated by differentially 
            targeted attachment molecule Num1.
  JOURNAL   Elife. 2018 Aug 7;7. pii: 36745. doi: 10.7554/eLife.36745.
  PUBMED    30084355
REFERENCE   2  (bases 1 to 5660)
  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: "Elife."; 
            date: "2018-08-7"; volume: "7. pii: 36745. doi"; pages: " 
            10.7554/eLife.36745"
COMMENT     Sequence Label: GFP-HDEL
FEATURES             Location/Qualifiers
     source          1..5660
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     17..34
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    151..172
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(187..192,193..210,211..217)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    225..241
                     /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     230..252
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     249..265
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     249..265
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        286..304
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     332..348
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     333..349
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(383..399)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(383..399)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     promoter        401..808
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     CDS             980..1687
                     /codon_start=1
                     /product="S65T variant of Aequorea victoria green 
                     fluorescent protein (Heim et al., 1995)"
                     /label=GFP (S65T)
                     /note="excitable with blue light"
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKF
                     ICTTGKLPVPWPTLVTTFTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTIFFKDDGN
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKVN
                     FKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELY"
     primer_bind     complement(1007..1035)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     1630..1650
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     primer_bind     complement(1722..1741)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1723..1741)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1748..1765)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1748..1764)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1757..1779)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      1905..2360
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1992..2011)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2202..2223
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2491..3294)
                     /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(3295..3510)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(3534..3553)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3653..3675
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3713..3731)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3799..3903
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3904..3972,3973..4764)
                     /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(4122..4141)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4935..5523
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5424..5443
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc caatacgcaa
       61 accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca ggtttcccga
      121 ctggaaagcg ggcagtgagc gcaacgcaat taatgtgagt tagctcactc attaggcacc
      181 ccaggcttta cactttatgc ttccggctcg tatgttgtgt ggaattgtga gcggataaca
      241 atttcacaca ggaaacagct atgaccatga ttacgccaag ctcgaaatta accctcacta
      301 aagggaacaa aagctggtac cgggcccccc ctcgaggtcg acggtatcga taagcttgat
      361 atcgaattcc tgcagcccgg gggatccact agttctagag ccacacacca tagcttcaaa
      421 atgtttctac tcctttttta ctcttccaga ttttctcgga ctccgcgcat cgccgtacca
      481 cttcaaaaca cccaagcaca gcatactaaa tttcccctct ttcttcctct agggtgtcgt
      541 taattacccg tactaaaggt ttggaaaaga aaaaagagac cgcctcgttt ctttttcttc
      601 gtcgaaaaag gcaataaaaa tttttatcac gtttcttttt cttgaaaatt tttttttttg
      661 atttttttct ctttcgatga cctcccattg atatttaagt taataaacgg tcttcaattt
      721 ctcaagtttc agtttcattt ttcttgttct attacaactt tttttacttc ttgctcatta
      781 gaaagaaagc atagcaatct aatctaagtt ttaattacaa atgtttttca acagactaag
      841 cgctggcaag ctgctggtac cactctccgt ggtcctgtac gcccttttcg tggtaatatt
      901 acctttacag aattctttcc actcctccaa tgttttagtt agaggtggtg ctggtgctgg
      961 tgctcccggg ttaattaaca gtaaaggaga agaacttttc actggagttg tcccaattct
     1021 tgttgaatta gatggtgatg ttaatgggca caaattttct gtcagtggag agggtgaagg
     1081 tgatgcaaca tacggaaaac ttacccttaa atttatttgc actactggaa aactacctgt
     1141 tccatggcca acacttgtca ctactttcac ttatggtgtt caatgctttt caagataccc
     1201 agatcatatg aaacggcatg actttttcaa gagtgccatg cccgaaggtt atgtacagga
     1261 aagaactata tttttcaaag atgacgggaa ctacaagaca cgtgctgaag tcaagtttga
     1321 aggtgatacc cttgttaata gaatcgagtt aaaaggtatt gattttaaag aagatggaaa
     1381 cattcttgga cacaaattgg aatacaacta taactcacac aatgtataca tcatggcaga
     1441 caaacaaaag aatggaatca aagttaactt caaaattaga cacaacattg aagatggaag
     1501 cgttcaacta gcagaccatt atcaacaaaa tactccaatt ggcgatggcc ctgtcctttt
     1561 accagacaac cattacctgt ccacacaatc tgccctttcg aaagatccca acgaaaagag
     1621 agaccacatg gtccttcttg agtttgtaac agctgctggg attacacatg gcatggatga
     1681 actataccac gacgaattgt aagcggtgga gctccaattc gccctatagt gagtcgtatt
     1741 acaattcact ggccgtcgtt ttacaacgtc gtgactggga aaaccctggc gttacccaac
     1801 ttaatcgcct tgcagcacat ccccctttcg ccagctggcg taatagcgaa gaggcccgca
     1861 ccgatcgccc ttcccaacag ttgcgcagcc tgaatggcga atggacgcgc cctgtagcgg
     1921 cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc
     1981 cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc
     2041 ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct
     2101 cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac
     2161 ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac
     2221 tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat
     2281 ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa
     2341 aatattaacg cttacaattt cctgatgcgg tattttctcc ttacgcatct gtgcggtatt
     2401 tcacaccgca tagggtaata actgatataa ttaaattgaa gctctaattt gtgagtttag
     2461 tatacatgca tttacttata atacagtttt ttagttttgc tggccgcatc ttctcaaata
     2521 tgcttcccag cctgcttttc tgtaacgttc accctctacc ttagcatccc ttccctttgc
     2581 aaatagtcct cttccaacaa taataatgtc agatcctgta gagaccacat catccacggt
     2641 tctatactgt tgacccaatg cgtctccctt gtcatctaaa cccacaccgg gtgtcataat
     2701 caaccaatcg taaccttcat ctcttccacc catgtctctt tgagcaataa agccgataac
     2761 aaaatctttg tcgctcttcg caatgtcaac agtaccctta gtatattctc cagtagatag
     2821 ggagcccttg catgacaatt ctgctaacat caaaaggcct ctaggttcct ttgttacttc
     2881 ttctgccgcc tgcttcaaac cgctaacaat acctgggccc accacaccgt gtgcattcgt
     2941 aatgtctgcc cattctgcta ttctgtatac acccgcagag tactgcaatt tgactgtatt
     3001 accaatgtca gcaaattttc tgtcttcgaa gagtaaaaaa ttgtacttgg cggataatgc
     3061 ctttagcggc ttaactgtgc cctccatgga aaaatcagtc aagatatcca catgtgtttt
     3121 tagtaaacaa attttgggac ctaatgcttc aactaactcc agtaattcct tggtggtacg
     3181 aacatccaat gaagcacaca agtttgtttg cttttcgtgc atgatattaa atagcttggc
     3241 agcaacagga ctaggatgag tagcagcacg ttccttatat gtagctttcg acatgattta
     3301 tcttcgtttc ctgcaggttt ttgttctgtg cagttgggtt aagaatactg ggcaatttca
     3361 tgtttcttca acactacata tgcgtatata taccaatcta agtctgtgct ccttccttcg
     3421 ttcttccttc tgttcggaga ttaccgaatc aaaaaaattt caaggaaacc gaaatcaaaa
     3481 aaaagaataa aaaaaaaatg atgaattgaa aaggtggtat ggtgcactct cagtacaatc
     3541 tgctctgatg ccgcatagtt aagccagccc cgacacccgc caacacccgc tgacgcgccc
     3601 tgacgggctt gtctgctccc ggcatccgct tacagacaag ctgtgaccgt ctccgggagc
     3661 tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg cgagacgaaa gggcctcgtg
     3721 atacgcctat ttttataggt taatgtcatg ataataatgg tttcttagac gtcaggtggc
     3781 acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat
     3841 atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag
     3901 agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt
     3961 cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt
     4021 gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga gagttttcgc
     4081 cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta
     4141 tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac
     4201 ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa
     4261 ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg
     4321 atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc
     4381 cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg
     4441 atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta
     4501 gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg accacttctg
     4561 cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg
     4621 tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc
     4681 tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt
     4741 gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt
     4801 gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc
     4861 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
     4921 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     4981 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     5041 aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag
     5101 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     5161 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     5221 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     5281 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     5341 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
     5401 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     5461 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     5521 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
     5581 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga
     5641 gctgataccg ctcgccgcag
//
