LOCUS       Exported                6002 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    pFRT/FLAG/HA-DEST LIN28A
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6002)
  TITLE     Identification of mRNAs bound and regulated by human LIN28 proteins 
            and molecular requirements for RNA recognition
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6002)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6002
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     CDS             1..42
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(86..105)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        277..656
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        657..860
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     811..831
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    862..880
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    883..901
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     911..935
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             1035..1058
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             1059..1085
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     protein_bind    1101..1125
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(1756..1780)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             1833..1874
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(1954..1971)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1960..2184
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2317..2336)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    2468..2515
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             2523..3545
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     polyA_signal    3675..3796
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3712..3731)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3766..3785
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3845..3861)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3845..3861)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3858..3880)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3869..3885
                     /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(3893..3923)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3938..3959
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4076..4093)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4247..4835)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4327..4346)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5006..5866)
                     /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     5629..5648
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5867..5971)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ggtaagccta tccctaaccc tctcctcggt ctcgattcta cggacggatc gggagatctc
       61 ccgatcccct atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagt
      121 atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg cgcgagcaaa atttaagcta
      181 caacaaggca aggcttgacc gacaattgca tgaagaatct gcttagggtt aggcgttttg
      241 cgctgcttcg cgatgtacgg gccagatata cgcgttgaca ttgattattg actagttatt
      301 aatagtaatc aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat
      361 aacttacggt aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa
      421 taatgacgta tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg
      481 agtatttacg gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc
      541 cccctattga cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct
      601 tatgggactt tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga
      661 tgcggttttg gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa
      721 gtctccaccc cattgacgtc aatgggagtt tgttttggaa ccaaaatcaa cgggactttc
      781 caaaatgtcg taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg
      841 aggtctatat aagcagagct ctccctatca gtgatagaga tctccctatc agtgatagag
      901 atcgtcgacg agctcgttta gtgaaccgtc agatcgcctg gagacgccat ccacgctgtt
      961 ttgacctcca tagaagacac cgggaccgat ccagcctccg gactctagcg tttaaactta
     1021 agcttggtac catggactac aaggacgacg atgacaagta cccttatgac gtgcccgatt
     1081 acgctttgaa ccagatatca acaagtttgt acaaaaaagc aggctatggg ctccgtgtcc
     1141 aaccagcagt ttgcaggtgg ctgcgccaag gcggcagaag aggcgcccga ggaggcgccg
     1201 gaggacgcgg cccgggcggc ggacgagcct cagctgctgc acggtgcggg catctgtaag
     1261 tggttcaacg tgcgcatggg gttcggcttc ctgtccatga ccgcccgcgc cggggtcgcg
     1321 ctcgaccccc cagtggatgt ctttgtgcac cagagtaagc tgcacatgga agggttccgg
     1381 agcttgaagg agggtgaggc agtggagttc acctttaaga agtcagccaa gggtctggaa
     1441 tccatccgtg tcaccggacc tggtggagta ttctgtattg ggagtgagag gcggccaaaa
     1501 ggaaagagca tgcagaagcg cagatcaaaa ggagacaggt gctacaactg tggaggtcta
     1561 gatcatcatg ccaaggaatg caagctgcca ccccagccca agaagtgcca cttctgccag
     1621 agcatcagcc atatggtagc ctcatgtccg ctgaaggccc agcagggccc tagtgcacag
     1681 ggaaagccaa cctactttcg agaggaagaa gaagaaatcc acagccctac cctgctcccg
     1741 gaggcacaga attgaaccca gctttcttgt acaaagtggt tgatatccag cacagtggcg
     1801 gccgctcaag tctagagggc ccgcggttcg aaggtaagcc tatccctaac cctctcctcg
     1861 gtctcgattc tacgcgtacc ggatccgcgg ccgcatagat aactgatcca gtgtgcctcg
     1921 agtctagagg gcccgtttaa acccgctgat cagcctcgac tgtgccttct agttgccagc
     1981 catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg
     2041 tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc
     2101 tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagacaat agcaggcatg
     2161 ctggggatgc ggtgggctct atggcttctg aggcggaaag aaccagctgg ggctctaggg
     2221 ggtatcccca cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca
     2281 gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct
     2341 ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt
     2401 tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac
     2461 gtacctagaa gttcctattc cgaagttcct attctctaga aagtatagga acttccttgg
     2521 ccaaaaagcc tgaactcacc gcgacgtctg tcgagaagtt tctgatcgaa aagttcgaca
     2581 gcgtctccga cctgatgcag ctctcggagg gcgaagaatc tcgtgctttc agcttcgatg
     2641 taggagggcg tggatatgtc ctgcgggtaa atagctgcgc cgatggtttc tacaaagatc
     2701 gttatgttta tcggcacttt gcatcggccg cgctcccgat tccggaagtg cttgacattg
     2761 gggaattcag cgagagcctg acctattgca tctcccgccg tgcacagggt gtcacgttgc
     2821 aagacctgcc tgaaaccgaa ctgcccgctg ttctgcagcc ggtcgcggag gccatggatg
     2881 cgatcgctgc ggccgatctt agccagacga gcgggttcgg cccattcgga ccgcaaggaa
     2941 tcggtcaata cactacatgg cgtgatttca tatgcgcgat tgctgatccc catgtgtatc
     3001 actggcaaac tgtgatggac gacaccgtca gtgcgtccgt cgcgcaggct ctcgatgagc
     3061 tgatgctttg ggccgaggac tgccccgaag tccggcacct cgtgcacgcg gatttcggct
     3121 ccaacaatgt cctgacggac aatggccgca taacagcggt cattgactgg agcgaggcga
     3181 tgttcgggga ttcccaatac gaggtcgcca acatcttctt ctggaggccg tggttggctt
     3241 gtatggagca gcagacgcgc tacttcgagc ggaggcatcc ggagcttgca ggatcgccgc
     3301 ggctccgggc gtatatgctc cgcattggtc ttgaccaact ctatcagagc ttggttgacg
     3361 gcaatttcga tgatgcagct tgggcgcagg gtcgatgcga cgcaatcgtc cgatccggag
     3421 ccgggactgt cgggcgtaca caaatcgccc gcagaagcgc ggccgtctgg accgatggct
     3481 gtgtagaagt actcgccgat agtggaaacc gacgccccag cactcgtccg agggcaaagg
     3541 aatagcacgt actacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg
     3601 gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt
     3661 tcttcgccca ccccaacttg tttattgcag cttataatgg ttacaaataa agcaatagca
     3721 tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac
     3781 tcatcaatgt atcttatcat gtctgtatac cgtcgacctc tagctagagc ttggcgtaat
     3841 catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac
     3901 gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa
     3961 ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat
     4021 gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc
     4081 tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg
     4141 cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag
     4201 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     4261 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     4321 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     4381 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     4441 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     4501 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     4561 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     4621 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     4681 ctagaaggac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     4741 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     4801 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     4861 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     4921 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     4981 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
     5041 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     5101 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
     5161 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
     5221 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
     5281 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
     5341 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
     5401 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
     5461 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
     5521 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
     5581 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
     5641 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
     5701 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
     5761 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
     5821 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
     5881 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     5941 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg
     6001 tc
//
