LOCUS       sequence-172605-        8620 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  luminal split GFP11 reporter with mCherry.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8620)
  AUTHORS   Smoyer CJ, Katta SS, Gardner JM, Stoltz L, McCroskey S, Bradford WD,
            McClain M, Smith SE, Slaughter BD, Unruh JR, Jaspersen SL
  TITLE     Analysis of membrane proteins localizing to the inner nuclear 
            envelope in living cells.
  JOURNAL   J Cell Biol. 2016 Nov 21;215(4):575-590. Epub 2016 Nov 9.
  PUBMED    27831485
REFERENCE   2  (bases 1 to 8620)
  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: "2016-11-21"; volume: "215(4)"; pages: "575-590. Epub 
            2016 Nov 9"
COMMENT     Sequence Label: pSJ1602 (pRS315-NOP1pr-mCherry-SCS2TM-GFP11)
FEATURES             Location/Qualifiers
     source          1..8620
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(297..885)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(377..396)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1056..1847,1848..1916))
                     /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     1679..1698
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1917..2021)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2089..2107
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(2145..2167)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2480..2502
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2494..2511
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2495..2511
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2542..2558
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2542..2558
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     CDS             4029..4076
                     /codon_start=1
                     /product="11th beta-strand of superfolder GFP"
                     /label=GFP11
                     /note="associates with GFP(1-10), consisting of the first 
                     10 beta-strands, to reconstitute fluorescent GFP (Cabantous
                     et al., 2005; Kamiyama et al., 2016)"
                     /translation="RDHMVLHEYVNAAGIT"
     primer_bind     complement(4104..4120)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4104..4120)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4117..4139)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4128..4144
                     /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(4152..4158,4159..4176,4177..4182))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4197..4218
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        4284..4659
                     /gene="S. cerevisiae ADE2"
                     /label=ADE2 promoter
     promoter        4851..5066
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     CDS             complement(5630..6724)
                     /codon_start=1
                     /gene="S. cerevisiae LEU2"
                     /product="3-isopropylmalate dehydrogenase, required for 
                     leucine biosynthesis"
                     /label=LEU2
                     /note="yeast auxotrophic marker"
                     /translation="MSAPKKIVVLPGDHVGQEITAEAIKVLKAISDVRSNVKFDFENHL
                     IGGAAIDATGVPLPDEALEASKKADAVLLGAVGGPKWGTGSVRPEQGLLKIRKELQLYA
                     NLRPCNFASDSLLDLSPIKPQFAKGTDFVVVRELVGGIYFGKRKEDDGDGVAWDSEQYT
                     VPEVQRITRMAAFMALQHEPPLPIWSLDKANVLASSRLWRKTVEETIKNEFPTLKVQHQ
                     LIDSAAMILVKNPTHLNGIIITSNMFGDIISDEASVIPGSLGLLPSASLASLPDKNTAF
                     GLYEPCHGSAPDLPKNKVNPIATILSAAMMSKLSLNLPEEGKAIEDAVKKVLDAGIRTG
                     DLGGSNSTAEVGDAVAEEVKKILA"
     promoter        complement(6725..7132)
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     primer_bind     complement(7918..7935)
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 tatatatata tatatatata aataataaaa acggaaagga aatagaataa taataataat
       61 actacacgtt tacgtatgta actcattcta ttattcagac tttgatatta atagaattat
      121 tactactagg tttcctcact tcaaaatcta taaatatacc gtcatgctct aatcgatgct
      181 cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg
      241 tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc
      301 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga
      361 aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct
      421 cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg
      481 gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag
      541 ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat
      601 cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac
      661 aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac
      721 tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc
      781 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
      841 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc
      901 ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg
      961 agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca
     1021 atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca
     1081 cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag
     1141 ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac
     1201 ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc
     1261 agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct
     1321 agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc
     1381 gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg
     1441 cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc
     1501 gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat
     1561 tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag
     1621 tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat
     1681 aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg
     1741 cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca
     1801 cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga
     1861 aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc
     1921 ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata
     1981 tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg
     2041 ccacctgacg tctaagaaac cattattatc atgacattaa cctataaaaa taggcgtatc
     2101 acgaggccct ttcgtctcgc gcgtttcggt gatgacggtg aaaacctctg acacatgcag
     2161 ctcccggaga cggtcacagc ttgtctgtaa gcggatgccg ggagcagaca agcccgtcag
     2221 ggcgcgtcag cgggtgttgg cgggtgtcgg ggctggctta actatgcggc atcagagcag
     2281 attgtactga gagtgcacca tatgcggtgt gaaataccgc acagatgcgt aaggagaaaa
     2341 taccgcatca ggcgccattc gccattcagg ctgcgcaact gttgggaagg gcgatcggtg
     2401 cgggcctctt cgctattacg ccagctggcg aaagggggat gtgctgcaag gcgattaagt
     2461 tgggtaacgc cagggttttc ccagtcacga cgttgtaaaa cgacggccag tgaattcgag
     2521 ctccaccgcg gtggcggccg ctctagaact agtggatcca ttgagtcatc agcctcttct
     2581 tgaaaaaacg attctgaaat aaccggaact tgtttagatt cctttttgta ggttaatctt
     2641 atcctgtact tgctgatatt gtggttatta gcagagattt ttttcaaaac atcatctaaa
     2701 gtaggctttt tggataagtc aatttcagtg tcccttaacc ctttagagcg gctttttatg
     2761 gtgataggca ttttcaaatt aaattcaaaa tatgtatctc tctcaaatag ctcaattggt
     2821 ttctagatag ctaaatagaa tataatctta ctgtcctccg ttctgtaaaa ttcacgctct
     2881 tagtcccttt tcataattcc ttaacttttt gcgtacaaaa tgatatgttt attatatttt
     2941 tctttttttt ttttcaaatt ttttcttttt cttgaaaaat ttttcaaatt ggaaagctca
     3001 tctctcttga atgtataata ctttcttcct ctaactttca aaaagtttta catagccaag
     3061 aagttttcct tacatcggta tactactgtt atataagtta ttcttcgaga aacaattaga
     3121 tatcattcat cggataaatc taagttgccc attgctttca ataactccga tcaaattaac
     3181 tcaaatcaac taaaacagta gggcccggag gcctcgagat ggtttctaaa ggtgaagaag
     3241 ataatatggc tattattaaa gaatttatga gatttaaagt tcatatggaa ggttctgtta
     3301 atggtcatga atttgaaatt gaaggtgaag gtgaaggtag accatatgaa ggtactcaaa
     3361 ctgctaaatt gaaagttact aaaggtggtc cattgccatt tgcttgggat attttgtctc
     3421 cacaatttat gtatggttct aaagcttatg ttaaacatcc agctgatatt ccagattatt
     3481 tgaaattgtc ttttccagaa ggttttaaat gggaaagagt tatgaatttt gaagatggtg
     3541 gtgttgttac tgttactcaa gattcttctt tgcaagatgg tgaatttatt tataaagtta
     3601 aattgagagg tactaatttt ccatctgatg gtccagttat gcaaaaaaaa actatgggtt
     3661 gggaagcttc ttctgaaaga atgtatccag aagatggtgc tttgaaaggt gaaattaaac
     3721 aaagattgaa attgaaagat ggtggtcatt atgatgctga agttaaaact acttataaag
     3781 ctaaaaaacc agttcaattg ccaggtgctt ataatgttaa tattaaattg gatattactt
     3841 ctcataatga agattatact attgttgaac aatatgaaag agctgaaggt agacattcta
     3901 ctggtggtat ggatgaattg tataaaggtg gtggtgctag ctcatccagc atgggtatat
     3961 tcatattggt tgcactcctt atcttggttt taggatggtt ctacagacca tggggtggtg
     4021 gttctatgag agatcatatg gttttgcatg aatatgttaa tgctgctggt attacttaag
     4081 catgcaagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc
     4141 acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga
     4201 gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg
     4261 tcgtgccagc agatctgaat taattcttga ataatacata acttttctta aaagaatcaa
     4321 agacagataa aatttaagag atattaaata ttagtgagaa gccgagaatt ttgtaacacc
     4381 aacataacac tgacatcttt aacaactttt aattatgata catttcttac gtcatgattg
     4441 attattacag ctatgctgac aaatgactct tgttgcatgg ctacgaaccg ggtaatacta
     4501 agtgattgac tcttgctgac cttttattaa gaactaaatg gacaatatta tggagcattt
     4561 catgtataaa ttggtgcgta aaatcgttgg atctctcttc taagtacatc ctactataac
     4621 aatcaagaaa aacaagaaaa tcggacaaaa caatcaagta tggattctag aacagttggt
     4681 atattaggag ggggacaatt gggacgtatg attgttgagg cagcaaacag gctcaacatt
     4741 aagacggtaa tactagatgc tgaaaattct cctgccaaac aaataagcaa ctccaatgac
     4801 cacgttaatg catcagagca gattgtactg agagtgcacc ataccacctt ttcaattcat
     4861 catttttttt ttattctttt ttttgatttc ggtttccttg aaattttttt gattcggtaa
     4921 tctccgaaca gaaggaagaa cgaaggaagg agcacagact tagattggta tatatacgca
     4981 tatgtagtgt tgaagaaaca tgaaattgcc cagtattctt aacccaactg cacagaacaa
     5041 aaacctgcag gaaacgaaga taaatcatgt cgaaagctac atataaggaa cgtgctgcta
     5101 ctcatcctag tcctgttgct gccaaggcag attgtactga gagtgcacca tatcgactac
     5161 gtcgttaagg ccgtttctga cagagtaaaa ttcttgaggg aactttcacc attatgggaa
     5221 atggttcaag aaggtattga cttaaactcc atcaaatggt caggtcattg agtgtttttt
     5281 atttgttgta tttttttttt tttagagaaa atcctccaat atataaatta ggaatcatag
     5341 tttcatgatt ttctgttaca cctaactttt tgtgtggtgc cctcctcctc gtcaatatta
     5401 atgttaaagt gcaattcttt ttccttatca cgttgagcca ttagtatcaa tttgcttacc
     5461 tgtattcctt tacatcctcc tttttctcct tcttgataaa tgtatgtaga ttgcgtatat
     5521 agtttcgtct accctatgaa catattccat tttgtaattt cgtgtcgttt ctattatgaa
     5581 tttcatttat aaagtttatg tacaaatatc ataaaaaaag agaatctttt taagcaagga
     5641 ttttcttaac ttcttcggcg acagcatcac cgacttcggc ggtactgttg gaaccaccta
     5701 aatcaccagt tctgatacct gcatccaaaa cctttttaac tgcatcttca atggccttac
     5761 cttcttcagg caagttcaat gacaatttcg acatcattgc agcagacaag atagtggcga
     5821 tagggttgac cttattcttt ggcaaatctg gagcagaacc gtggcatggt tcgtacaaac
     5881 caaatgcggt gttcttgtct ggcaaagagg ccaaggacgc agatggcaac aaacccaagg
     5941 aacctgggat aacggaggct tcatcggaga tgatatcacc aaacatgttg ctggtgatta
     6001 taataccatt taggtgggtt gggttcttaa ctaggatcat ggcggcagaa tcaatcaatt
     6061 gatgttgaac cttcaatgta gggaattcgt tcttgatggt ttcctccaca gtttttctcc
     6121 ataatcttga agaggccaaa acattagctt tatccaagga ccaaataggc aatggtggct
     6181 catgttgtag ggccatgaaa gcggccattc ttgtgattct ttgcacttct ggaacggtgt
     6241 attgttcact atcccaagcg acaccatcac catcgtcttc ctttctctta ccaaagtaaa
     6301 tacctcccac taattctctg acaacaacga agtcagtacc tttagcaaat tgtggcttga
     6361 ttggagataa gtctaaaaga gagtcggatg caaagttaca tggtcttaag ttggcgtaca
     6421 attgaagttc tttacggatt tttagtaaac cttgttcagg tctaacacta ccggtacccc
     6481 atttaggacc acccacagca cctaacaaaa cggcatcagc cttcttggag gcttccagcg
     6541 cctcatctgg aagtggaaca cctgtagcat cgatagcagc accaccaatt aaatgatttt
     6601 cgaaatcgaa cttgacattg gaacgaacat cagaaatagc tttaagaacc ttaatggctt
     6661 cggctgtgat ttcttgacca acgtggtcac ctggcaaaac gacgatcttc ttaggggcag
     6721 acattagaat ggtatatcct tgaaatatat atatatatat tgctgaaatg taaaaggtaa
     6781 gaaaagttag aaagtaagac gattgctaac cacctattgg aaaaaacaat aggtccttaa
     6841 ataatattgt caacttcaag tattgtgatg caagcattta gtcatgaacg cttctctatt
     6901 ctatatgaaa agccggttcc ggcgctctca cctttccttt ttctcccaat ttttcagttg
     6961 aaaaaggtat atgcgtcagg cgacctctga aattaacaaa aaatttccag tcatcgaatt
     7021 tgattctgtg cgatagcgcc cctgtgtgtt ctcgttatgt tgaggaaaaa aataatggtt
     7081 gctaagagat tcgaactctt gcatcttacg atacctgagt attcccacag ttaactgcgg
     7141 tcaagatatt tcttgaatca ggcgccttag accgctcggc caaacaacca attacttgtt
     7201 gagaaataga gtataattat cctataaata taacgctttt gaacacacat gaacagggaa
     7261 gtacaggaca attgattttg aagagaatgt ggattttgat gtaattgttg ggattccatt
     7321 tttaataagg caataatatt aggtatgtag atatactaga agttctcctc gaccggtcga
     7381 tatgcggtgt gaaataccgc acagagggtg aacgttacag aaaagcaggc tgggaagcat
     7441 atttgagaag atgcggccag caaaactaaa aaactgtatt ataagtaaat gcatgtatac
     7501 taaactcaca aattagagct tcaatttaat tatatcagtt attaccctat gcggtgtgaa
     7561 ataccgcaca gatgcgtagg agattcaagt tatacaacga aaatggaaca gtttttatta
     7621 aagcaagaag aagaagttct tgtcaaagca caaaagttag aaactgtcgg ttacgaagct
     7681 tatctagaaa acaagtaata tataagttta ttgatatact tgtacagcaa ataattataa
     7741 aatgatatac ctatttttta ggctttgtta tgattacatc aaatgtggac ttcatacata
     7801 gaaatcaacg cttacaggtg tcctttttta agaatttcat acataagatc tgctgcatta
     7861 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc
     7921 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag catcgatgct
     7981 agagtcgaat ggaaattacg tactttcaac gcaagagcaa gacaaaagaa agcctaaaac
     8041 tgtcttttcg tgtcggtcgt ccaatatcat cgtaaacgtg tgtagagcct taaattctaa
     8101 tttctaactt taaatttttt ttgctattat aatactaatt tctaactttc aacctatttt
     8161 acatcttcgg aaaacaaaaa ctattttttc tttaatttct ttttttactt tctattttta
     8221 atttatatat ttatattaaa aaatttaaat tataattatt tttatagcac gtgatgaaaa
     8281 gaaatatagt cttcaaggtt atattataca ggtttatata ttattttact tatttttgtt
     8341 gttctttttc ttggagtatt tagcattaac aacttcgacc catcgcgatg acgaaattgc
     8401 acgtttaaag gagaagtcgg aatcgttgca agaagaaata aattagtttg acgttttatt
     8461 ccttttcttc tgcacagaag tgtgtatata tatatttata tatagtcgat gtgcaaaagc
     8521 ctacattatt tttcttttga tttttatttt tttgttcaag taggttcatg ccgacagtaa
     8581 atccttatgc cctacctaaa tataaacacc caaatatata
//
