LOCUS       sequence-12727-e        4673 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 4673)
  AUTHORS   Tagwerker C, Zhang H, Wang X, Larsen LS, Lathrop RH, Hatfield GW, 
            Auer B, Huang L, Kaiser P
  TITLE     HB tag modules for PCR-based gene tagging and tandem affinity 
            purification in Saccharomyces cerevisiae.
  JOURNAL   Yeast. 2006 Jun . 23(8):623-32.
  PUBMED    16823883
REFERENCE   2  (bases 1 to 4673)
  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: "Yeast. 2006
            Jun . 23(8):623-32."
COMMENT     Sequence Label: pFA6a-HBH-hphMX4
FEATURES             Location/Qualifiers
     source          1..4673
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             72..89
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             318..335
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      362..549
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(417..441)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     gene            624..2199
                     /label=hphMX6
                     /note="yeast selectable marker conferring hygromycin 
                     resistance"
     promoter        624..967
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             968..1996
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     CDS             968..1996
                     /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      2002..2199
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(2310..2329)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2311..2329)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2416..2433)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2587..3175)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2667..2686)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3346..4137,4138..4206))
                     /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     3969..3988
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4207..4311)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4379..4397
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4435..4457)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4557..4576
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        join(4657..4673,1..2)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     join(4657..4673,1)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
ORIGIN
        1 gaacgcggcc gccagctgaa gcttcgtacg ctgcaggtcg acggatcccc gggttaatta
       61 acaggggttc acatcatcac caccatcatg ctggaaaggc cggtgaaggt gaaatccctg
      121 cccctcttgc tggtaccgtt tctaagatac tggtaaaaga aggtgacact gttaaagctg
      181 gtcaaacagt tctggtgctg gaggctatga aaatggagac agaaattaac gctcctactg
      241 acggaaaagt tgaaaaggtg ttagttaagg aaagagatgc tgttcaaggt ggtcaaggtc
      301 taatcaagat cggcgttcat catcaccacc atcattaatg aggcgcgcca cttctaaata
      361 agcgaatttc ttatgattta tgatttttat tattaaataa gttataaaaa aaataagtgt
      421 atacaaattt taaagtgact cttaggtttt aaaacgaaaa ttcttattct tgagtaactc
      481 tttcctgtag gtcaggttgc tttctcaggt atagtatgag gtcgctctta ttgaccacac
      541 ctctaccggc agatccgcta gggataacag ggtaatatag atctgtttag cttgccttgt
      601 ccccgccggg tcacccggcc agcgacatgg aggcccagaa taccctcctt gacagtcttg
      661 acgtgcgcag ctcaggggca tgatgtgact gtcgcccgta catttagccc atacatcccc
      721 atgtataatc atttgcatcc atacattttg atggccgcac ggcgcgaagc aaaaattacg
      781 gctcctcgct gcagacctgc gagcagggaa acgctcccct cacagacgcg ttgaattgtc
      841 cccacgccgc gcccctgtag agaaatataa aaggttagga tttgccactg aggttcttct
      901 ttcatatact tccttttaaa atcttgctag gatacagttc tcacatcaca tccgaacata
      961 aacaaccatg ggtaaaaagc ctgaactcac cgcgacgtct gtcgagaagt ttctgatcga
     1021 aaagttcgac agcgtctccg acctgatgca gctctcggag ggcgaagaat ctcgtgcttt
     1081 cagcttcgat gtaggagggc gtggatatgt cctgcgggta aatagctgcg ccgatggttt
     1141 ctacaaagat cgttatgttt atcggcactt tgcatcggcc gcgctcccga ttccggaagt
     1201 gcttgacatt ggggaattca gcgagagcct gacctattgc atctcccgcc gtgcacaggg
     1261 tgtcacgttg caagacctgc ctgaaaccga actgcccgct gttctgcagc cggtcgcgga
     1321 ggccatggat gcgatcgctg cggccgatct tagccagacg agcgggttcg gcccattcgg
     1381 accgcaagga atcggtcaat acactacatg gcgtgatttc atatgcgcga ttgctgatcc
     1441 ccatgtgtat cactggcaaa ctgtgatgga cgacaccgtc agtgcgtccg tcgcgcaggc
     1501 tctcgatgag ctgatgcttt gggccgagga ctgccccgaa gtccggcacc tcgtgcacgc
     1561 ggatttcggc tccaacaatg tcctgacgga caatggccgc ataacagcgg tcattgactg
     1621 gagcgaggcg atgttcgggg attcccaata cgaggtcgcc aacatcttct tctggaggcc
     1681 gtggttggct tgtatggagc agcagacgcg ctacttcgag cggaggcatc cggagcttgc
     1741 aggatcgccg cggctccggg cgtatatgct ccgcattggt cttgaccaac tctatcagag
     1801 cttggttgac ggcaatttcg atgatgcagc ttgggcgcag ggtcgatgcg acgcaatcgt
     1861 ccgatccgga gccgggactg tcgggcgtac acaaatcgcc cgcagaagcg cggccgtctg
     1921 gaccgatggc tgtgtagaag tactcgccga tagtggaaac cgacgcccca gcactcgtcc
     1981 gagggcaaag gaataatcag tactgacaat aaaaagattc ttgttttcaa gaacttgtca
     2041 tttgtatagt ttttttatat tgtagttgtt ctattttaat caaatgttag cgtgatttat
     2101 attttttttc gcctcgacat catctgccca gatgcgaagt taagtgcgca gaaagtaata
     2161 tcatgcgtca atcgtatgtg aatgctggtc gctatactgc tgtcgattcg atactaacgc
     2221 cgccatccag tgtcgagttt aaacgagctc gaattcatcg atgatatcag atccactagt
     2281 ggcctatgcg gccgcggatc tgccggtctc cctatagtga gtcgtattaa tttcgataag
     2341 ccaggttaac ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg
     2401 gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc
     2461 ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg
     2521 aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct
     2581 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca
     2641 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
     2701 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
     2761 gggaagcgtg gcgctttctc aatgctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
     2821 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
     2881 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
     2941 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
     3001 gtggcctaac tacggctaca ctagaaggac agtatttggt atctgcgctc tgctgaagcc
     3061 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
     3121 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga
     3181 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat
     3241 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag
     3301 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat
     3361 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc
     3421 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat
     3481 accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag
     3541 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg
     3601 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc
     3661 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca
     3721 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg
     3781 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc
     3841 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta
     3901 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc
     3961 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg
     4021 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc
     4081 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc
     4141 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat
     4201 actcatactc ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag
     4261 cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc
     4321 ccgaaaagtg ccacctgacg tctaagaaac cattattatc atgacattaa cctataaaaa
     4381 taggcgtatc acgaggccct ttcgtctcgc gcgtttcggt gatgacggtg aaaacctctg
     4441 acacatgcag ctcccggaga cggtcacagc ttgtctgtaa gcggatgccg ggagcagaca
     4501 agcccgtcag ggcgcgtcag cgggtgttgg cgggtgtcgg ggctggctta actatgcggc
     4561 atcagagcag attgtactga gagtgcacca tatggacata ttgtcgttag aacgcggcta
     4621 caattaatac ataaccttat gtatcataca catacgattt aggtgacact ata
//
