LOCUS       sequence-61675-e        4482 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4482)
  TITLE     Formosa lab yeast 3' tagging constructs
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 4482)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pTF271 (pFA6a-TEV-6xGly-12xHis-HphMX)
FEATURES             Location/Qualifiers
     source          1..4482
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(199..303)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     371..389
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(427..449)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     549..568
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        649..667
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     649..666
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     CDS             728..748
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             776..805
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             779..808
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             782..811
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     terminator      840..1027
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(895..919)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     gene            1102..2677
                     /label=hphMX6
                     /note="yeast selectable marker conferring hygromycin 
                     resistance"
     promoter        1102..1445
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             1446..2474
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     CDS             1446..2474
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     terminator      2480..2677
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(2784..2803)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2785..2803)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2890..2907)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3061..3649)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3141..3160)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3820..4482,1..129,130..198))
                     /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     4443..4462
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
       61 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
      121 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
      181 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
      241 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
      301 gcgcacattt ccccgaaaag tgccacctga cgtctaagaa accattatta tcatgacatt
      361 aacctataaa aataggcgta tcacgaggcc ctttcgtctc gcgcgtttcg gtgatgacgg
      421 tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt aagcggatgc
      481 cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc ggggctggct
      541 taactatgcg gcatcagagc agattgtact gagagtgcac catatggaca tattgtcgtt
      601 agaacgcggc tacaattaat acataacctt atgtatcata cacatacgat ttaggtgaca
      661 ctatagaacg cggccgccag ctgaagcttc gtacgctgca ggtcgacgga tccccgggtt
      721 aattaacgag aatttatact tccaaggttc tggtggaggc ggtggaggct ctagacatca
      781 ccaccatcat caccaccatc atcatcacca ttgactcgag gcgcgccact tctaaataag
      841 cgaatttctt atgatttatg atttttatta ttaaataagt tataaaaaaa ataagtgtat
      901 acaaatttta aagtgactct taggttttaa aacgaaaatt cttattcttg agtaactctt
      961 tcctgtaggt caggttgctt tctcaggtat agtatgaggt cgctcttatt gaccacacct
     1021 ctaccggcag atccgctagg gataacaggg taatatagat ctgtttagct tgcctcgtcc
     1081 ccgccgggtc acccggccag cgacatggag gcccagaata ccctccttga cagtcttgac
     1141 gtgcgcagct caggggcatg atgtgactgt cgcccgtaca tttagcccat acatccccat
     1201 gtataatcat ttgcatccat acattttgat ggccgcacgg cgcgaagcaa aaattacggc
     1261 tcctcgctgc agacctgcga gcagggaaac gctcccctca cagacgcgtt gaattgtccc
     1321 cacgccgcgc ccctgtagag aaatataaaa ggttaggatt tgccactgag gttcttcttt
     1381 catatacttc cttttaaaat cttgctagga tacagttctc acatcacatc cgaacataaa
     1441 caaccatggg taaaaagcct gaactcaccg cgacgtctgt cgagaagttt ctgatcgaaa
     1501 agttcgacag cgtctccgac ctgatgcagc tctcggaggg cgaagaatct cgtgctttca
     1561 gcttcgatgt aggagggcgt ggatatgtcc tgcgggtaaa tagctgcgcc gatggtttct
     1621 acaaagatcg ttatgtttat cggcactttg catcggccgc gctcccgatt ccggaagtgc
     1681 ttgacattgg ggaattcagc gagagcctga cctattgcat ctcccgccgt gcacagggtg
     1741 tcacgttgca agacctgcct gaaaccgaac tgcccgctgt tctgcagccg gtcgcggagg
     1801 ccatggatgc gatcgctgcg gccgatctta gccagacgag cgggttcggc ccattcggac
     1861 cgcaaggaat cggtcaatac actacatggc gtgatttcat atgcgcgatt gctgatcccc
     1921 atgtgtatca ctggcaaact gtgatggacg acaccgtcag tgcgtccgtc gcgcaggctc
     1981 tcgatgagct gatgctttgg gccgaggact gccccgaagt ccggcacctc gtgcacgcgg
     2041 atttcggctc caacaatgtc ctgacggaca atggccgcat aacagcggtc attgactgga
     2101 gcgaggcgat gttcggggat tcccaatacg aggtcgccaa catcttcttc tggaggccgt
     2161 ggttggcttg tatggagcag cagacgcgct acttcgagcg gaggcatccg gagcttgcag
     2221 gatcgccgcg gctccgggcg tatatgctcc gcattggtct tgaccaactc tatcagagct
     2281 tggttgacgg caatttcgat gatgcagctt gggcgcaggg tcgatgcgac gcaatcgtcc
     2341 gatccggagc cgggactgtc gggcgtacac aaatcgcccg cagaagcgcg gccgtctgga
     2401 ccgatggctg tgtagaagta ctcgccgata gtggaaaccg acgccccagc actcgtccga
     2461 gggcaaagga ataatcagta ctgacaataa aaagattctt gttttcaaga acttgtcatt
     2521 tgtatagttt ttttatattg tagttgttct attttaatca aatgttagcg tgatttatat
     2581 tttttttcgc ctcgacatca tctgcccaga tgcgaagtta agtgcgcaga aagtaatatc
     2641 atgcgtcaat cgtatgtgaa tgctggtcgc tatactgctg tcgattcgat actaacgccg
     2701 ccatccagtg tcgaaaacga gctcgaattc atcgatgata tcagatccac tagtggccta
     2761 tgcggccgcg gatctgccgg tctccctata gtgagtcgta ttaatttcga taagccaggt
     2821 taacctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc
     2881 ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc
     2941 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa
     3001 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
     3061 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
     3121 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
     3181 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     3241 cgtggcgctt tctcaatgct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     3301 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
     3361 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     3421 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
     3481 taactacggc tacactagaa ggacagtatt tggtatctgc gctctgctga agccagttac
     3541 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
     3601 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
     3661 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
     3721 catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa
     3781 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga
     3841 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
     3901 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg
     3961 agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga
     4021 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
     4081 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg
     4141 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
     4201 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
     4261 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
     4321 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
     4381 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg
     4441 ggataatacc gcgccacata gcagaacttt aaaagtgctc at
//
