LOCUS       Exported                5570 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    SIRT3L-12
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5570)
  AUTHORS   Hallows WC, Lee S, Denu JM
  TITLE     Sirtuins deacetylate and activate mammalian acetyl-CoA synthetases.
  JOURNAL   Proc Natl Acad Sci U S A. 2006 Jul 5. 103(27):10230-5.
  PUBMED    16790548
REFERENCE   2  (bases 1 to 5570)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5570
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     promoter        10..54
                     /label=T5 promoter
                     /note="bacteriophage T5 promoter for E. coli RNA 
                     polymerase, with embedded lac operator"
     protein_bind    30..46
                     /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)."
     protein_bind    62..78
                     /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)."
     RBS             97..108
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             127..144
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      1027..1121
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             1165..1824
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(1231..1250)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      1889..1975
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(2068..3150)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     3112..3131
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(3151..3228)
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     primer_bind     complement(3312..3334)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3420..3560
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3475..3494
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3575..3592)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3746..4334)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3826..3845)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(4505..5365)
                     /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     5128..5147
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5366..5470)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5538..5556
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 ctcgagaaat cataaaaaat ttatttgctt tgtgagcgga taacaattat aatagattca
       61 attgtgagcg gataacaatt tcacacagaa ttcattaaag aggagaaatt aactatgaga
      121 ggatcgcatc accatcacca tcacggatcc aaggggaagc tttccctgca ggatgtagct
      181 gagctgattc gggccagagc ctgccagagg gtggtggtca tggtgggggc cggcatcagc
      241 acacccagtg gcattccaga cttcagatcg ccggggagtg gcctgtacag caacctccag
      301 cagtacgatc tcccgtaccc cgaggccatt tttgaactcc cattcttctt tcacaacccc
      361 aagccctttt tcactttggc caaggagctg taccctggaa actacaagcc caacgtcact
      421 cactactttc tccggctgct tcatgacaag gggctgcttc tgcggctcta cacgcagaac
      481 atcgatgggc ttgagagagt gtcgggcatc cctgcctcaa agctggttga agctcatgga
      541 acctttgcct ctgccacctg cacagtctgc caaagaccct tcccagggga ggacattcgg
      601 gctgacgtga tggcagacag ggttccccgc tgcccggtct gcaccggcgt tgtgaagccc
      661 gacattgtgt tctttgggga gccgctgccc cagaggttct tgctgcatgt ggttgatttc
      721 cccatggcag atctgctgct catccttggg acctccctgg aggtggagcc ttttgccagc
      781 ttgaccgagg ccgtgcggag ctcagttccc cgactgctca tcaaccggga cttggtgggg
      841 cccttggctt ggcatcctcg cagcagggac gtggcccagc tgggggacgt ggttcacggc
      901 gtggaaagcc tagtggagct tctgggctgg acagaagaga tgcgggacct tgtgcagcgg
      961 gaaactggga agcttgatgg accagacaaa tgtcgacctg cagccaagct taattagctg
     1021 agcttggact cctgttgata gatccagtaa tgacctcaga actccatctg gatttgttca
     1081 gaacgctcgg ttgccgccgg gcgtttttta ttggtgagaa tccaagctag cttggcgaga
     1141 ttttcaggag ctaaggaagc taaaatggag aaaaaaatca ctggatatac caccgttgat
     1201 atatcccaat ggcatcgtaa agaacatttt gaggcatttc agtcagttgc tcaatgtacc
     1261 tataaccaga ccgttcagct ggatattacg gcctttttaa agaccgtaaa gaaaaataag
     1321 cacaagtttt atccggcctt tattcacatt cttgcccgcc tgatgaatgc tcatccggaa
     1381 tttcgtatgg caatgaaaga cggtgagctg gtgatatggg atagtgttca cccttgttac
     1441 accgttttcc atgagcaaac tgaaacgttt tcatcgctct ggagtgaata ccacgacgat
     1501 ttccggcagt ttctacacat atattcgcaa gatgtggcgt gttacggtga aaacctggcc
     1561 tatttcccta aagggtttat tgagaatatg tttttcgtct cagccaatcc ctgggtgagt
     1621 ttcaccagtt ttgatttaaa cgtggccaat atggacaact tcttcgcccc cgttttcacc
     1681 atgggcaaat attatacgca aggcgacaag gtgctgatgc cgctggcgat tcaggttcat
     1741 catgccgttt gtgatggctt ccatgtcggc agaatgctta atgaattaca acagtactgc
     1801 gatgagtggc agggcggggc gtaatttttt taaggcagtt attggtgccc ttaaacgcct
     1861 ggggtaatga ctctctagct tgaggcatca aataaaacga aaggctcagt cgaaagactg
     1921 ggcctttcgt tttatctgtt gtttgtcggt gaacgctctc ctgagtagga caaatccgcc
     1981 ctctagatta cgtgcagtcg atgataagct gtcaaacatg agaattgtgc ctaatgagtg
     2041 agctaactta cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg
     2101 tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc
     2161 cagggtggtt tttcttttca ccagtgagac gggcaacagc tgattgccct tcaccgcctg
     2221 gccctgagag agttgcagca agcggtccac gctggtttgc cccagcaggc gaaaatcctg
     2281 tttgatggtg gttaacggcg ggatataaca tgagctgtct tcggtatcgt cgtatcccac
     2341 taccgagata tccgcaccaa cgcgcagccc ggactcggta atggcgcgca ttgcgcccag
     2401 cgccatctga tcgttggcaa ccagcatcgc agtgggaacg atgccctcat tcagcatttg
     2461 catggtttgt tgaaaaccgg acatggcact ccagtcgcct tcccgttccg ctatcggctg
     2521 aatttgattg cgagtgagat atttatgcca gccagccaga cgcagacgcg ccgagacaga
     2581 acttaatggg cccgctaaca gcgcgatttg ctggtgaccc aatgcgacca gatgctccac
     2641 gcccagtcgc gtaccgtctt catgggagaa aataatactg ttgatgggtg tctggtcaga
     2701 gacatcaaga aataacgccg gaacattagt gcaggcagct tccacagcaa tggcatcctg
     2761 gtcatccagc ggatagttaa tgatcagccc actgacgcgt tgcgcgagaa gattgtgcac
     2821 cgccgcttta caggcttcga cgccgcttcg ttctaccatc gacaccacca cgctggcacc
     2881 cagttgatcg gcgcgagatt taatcgccgc gacaatttgc gacggcgcgt gcagggccag
     2941 actggaggtg gcaacgccaa tcagcaacga ctgtttgccc gccagttgtt gtgccacgcg
     3001 gttgggaatg taattcagct ccgccatcgc cgcttccact ttttcccgcg ttttcgcaga
     3061 aacgtggctg gcctggttca ccacgcggga aacggtctga taagagacac cggcatactc
     3121 tgcgacatcg tataacgtta ctggtttcac attcaccacc ctgaattgac tctcttccgg
     3181 gcgctatcat gccataccgc gaaaggtttt gcaccattcg atggtgtcgg aatttcgggc
     3241 agcgttgggt cctggccacg ggtgcgcatg atctagagct gcctcgcgcg tttcggtgat
     3301 gacggtgaaa acctctgaca catgcagctc ccggagacgg tcacagcttg tctgtaagcg
     3361 gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc
     3421 gcagccatga cccagtcacg tagcgatagc ggagtgtata ctggcttaac tatgcggcat
     3481 cagagcagat tgtactgaga gtgcaccata tgcggtgtga aataccgcac agatgcgtaa
     3541 ggagaaaata ccgcatcagg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
     3601 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     3661 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     3721 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     3781 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     3841 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     3901 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     3961 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     4021 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     4081 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     4141 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaaggaca gtatttggta
     4201 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     4261 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     4321 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     4381 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     4441 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
     4501 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
     4561 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg
     4621 gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa
     4681 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca
     4741 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc
     4801 gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt
     4861 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa
     4921 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat
     4981 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct
     5041 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga
     5101 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag
     5161 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga
     5221 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca
     5281 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg
     5341 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc
     5401 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
     5461 gggttccgcg cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc attattatca
     5521 tgacattaac ctataaaaat aggcgtatca cgaggccctt tcgtcttcac
//
