LOCUS       Exported                5715 bp DNA     circular SYN 02-JUL-2021
DEFINITION  BBF10K_000518 in pOpen_v4.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5715)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jul 2, 2021 from SnapGene 5.3.1
            https://www.snapgene.com
COMMENT     This file was created using tools provided by Twist Bioscience
FEATURES             Location/Qualifiers
     source          1..5715
                     /organism="BBF10K_000518 in pOpen_v4"
     primer_bind     21..51
                     /label=Gibson-for
     CDS             52..1221
                     /codon_start=1
                     /product="transforming growth factor beta 1"
                     /label=TGFb
                     /translation="MPPSGLRLLPLLLPLLWLLVLTPGRPAAGLSTCKTIDMELVKRKR
                     IEAIRGQILSKLRLASPPSQGEVPPGPLPEAVLALYNSTRDRVAGESAEPEPEPEADYY
                     AKEVTRVLMVETHNEIYDKFKQSTHSIYMFFNTSELREAVPEPVLLSRAELRLLRLKLK
                     VEQHVELYQKYSNNSWRYLSNRLLAPSDSPEWLSFDVTGVVRQWLSRGGEIEGFRLSAH
                     CSCDSRDNTLQVDINGFTTGRRGDLATIHGMNRPFLLLMATPLERAQHLQSSRHRRALD
                     TNYCFSSTEKNCCVRQLYIDFRKDLGWKWIHEPKGYHANFCLGPCPYIWSLDTQYSKVL
                     ALYNQHNPGASAAPCCVPQALEPLPIVYYVGRKPKVEQLSNMIVRSCKCS"
     primer_bind     complement(1222..1246)
                     /label=Gibson-rev
     terminator      1365..1412
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(1417..1433)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    1592..1842
                     /gene="incC"
                     /label=incC
                     /note="incompatibility region of the bacterial F plasmid"
     CDS             complement(1845..2600)
                     /codon_start=1
                     /gene="repE"
                     /product="replication initiation protein for the bacterial 
                     F plasmid"
                     /label=repE
                     /translation="MAETAVINHKKRKNSPRIVQSNDLTEAAYSLSRDQKRMLYLFVDQ
                     IRKSDGTLQEHDGICEIHVAKYAEIFGLTSAEASKDIRQALKSFAGKEVVFYRPEEDAG
                     DEKGYESFPWFIKRAHSPSRGLYSVHINPYLIPFFIGLQNRFTQFRLSETKEITNPYAM
                     RLYESLCQYRKPDGSGIVSLKIDWIIERYQLPQSYQRMPDFRRRFLQVCVNEINSRTPM
                     RLSYIEKKKGRQTTHIVFSFRDITSMTTG"
     rep_origin      2691..2910
                     /label=ori2
                     /note="secondary origin of replication for the bacterial F 
                     plasmid; also known as oriS"
     protein_bind    3330..3351
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        3366..3396
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3404..3420
                     /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)."
     primer_bind     3428..3444
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             3517..4362
                     /codon_start=1
                     /label=repL
                     /translation="MLAKVTFLSCITMSDFTFSGYELACFVTHSGLSRSAGHILSQCAN
                     LAATTSEYFIHKPHRLIAAETGYSQSTVVRAFREAVNKGILSVEIVIGDHRERRANLYR
                     FTPSFLAFAQQAKNALIESKLKISSAATKVKAVLAKTLALFNFLSTPPCQNDTPSPCQD
                     DVAIKNKKSQVKKTKRSVSGGAGTTSLKKLTSWIAKAKAKADNLRLSKKRTQKHEFKQK
                     VEAAARKYAYLKNKRSPDIGGISNFDNLPHCMTVNEALNAVLAKNKDNEQWGIPAGFRG
                     "
     misc_feature    4470..4621
                     /label=nnpp promoters
     CDS             4661..5476
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     terminator      complement(5586..5629)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     primer_bind     5647..5663
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     terminator      5664..5695
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
ORIGIN
        1 aagaattact gacccctcgg cagactgagt cgcccggggg gatccgccac catgcccccc
       61 tcaggactcc gactcctgcc tctccttctt cctcttcttt ggcttcttgt tcttactcct
      121 ggacggcctg cagctggtct tagtacttgt aaaacaattg atatggaact cgtcaaaagg
      181 aaacgaattg aagcgattcg cggacaaatt ctttctaaat tgcgcctcgc ttccccacct
      241 tcacaagggg aagttccacc tgggcctctt ccagaagcag tactcgcact ttataattct
      301 acacgcgatc gggttgctgg agaatcagca gaacctgaac cagaaccaga agcagattat
      361 tatgctaaag aagtaacccg agttttgatg gttgagacac ataatgagat atacgataaa
      421 tttaagcaat ccacacattc catctatatg ttctttaata cttcagaact ccgggaagct
      481 gtaccggaac ctgtcttgct cagtcgtgca gaacttcgcc tgctccgttt gaaacttaaa
      541 gtcgaacaac atgtcgaact ctatcagaaa tattcaaata atagttggcg ctatctctct
      601 aatcgactcc tcgcaccttc agattcccct gaatggcttt catttgatgt cacaggtgtc
      661 gtaaggcaat ggctttcacg gggcggagaa attgaagggt ttaggcttag tgctcattgt
      721 agttgtgata gtcgggataa tacattgcaa gtagatatta atggatttac aactgggcgt
      781 aggggagatc tcgctactat tcatggaatg aataggcctt ttctcttgct catggctact
      841 ccacttgaac gagcgcaaca tcttcaatca tcacggcatc ggcgtgctct tgatactaat
      901 tattgtttct cctctactga gaagaattgt tgcgtcaggc aattgtatat tgattttagg
      961 aaagatctcg ggtggaaatg gattcatgag cctaaaggat atcacgctaa cttttgtctt
     1021 ggtccttgtc catatatatg gtccctcgat acacaatatt ccaaagtctt ggctttgtat
     1081 aatcaacata atcctggggc atccgcggca ccatgttgtg tacctcaagc attggaacca
     1141 ctccctatag tctattatgt tgggcgaaag cccaaggtgg agcagctgag caacatgatc
     1201 gtgagaagct gcaagtgcag cggatccgcc accaacttca gcctgctaga ttagctgccg
     1261 atactggtgc gttacgcatg atacttaata gggataacag ggtaatgggt ccacaagatt
     1321 ggaacgtcgc tgaggtgtca atcgtcggag ccgctgagca ataactagca taaccccttg
     1381 gggcctctaa acgggtcttg aggggttttt tgcatggtca tagctgtttc ctgagagctt
     1441 ggcaggtgtc gcgtatgcat cgatcacgtg tccacgttct ttaatagtgg actcttgttc
     1501 caaactggaa caacactcgg atcgatccgg cgcgcaccgt gggaaaaact ccaggtagag
     1561 gtacacacgc ggatagccaa ttcagagtaa taaactgtga taatcaaccc tcatcaatga
     1621 tgacgaacta acccccgata tcaggtcaca tgacgaaggg aaagagaagg aaatcaactg
     1681 tgacaaactg ccctcaaatt tggcttcctt aaaaattaca gttcaaaaag tatgagaaaa
     1741 tccatgcagg ctgaaggaaa cagcaaaact gtgacaaatt accctcagta ggtcagaaca
     1801 aatgtgacga accaccctca aatctgtgac agataaccct cagactatcc tgtcgtcatg
     1861 gaagtgatat cgcggaagga aaatacgata tgagtcgtct ggcggccttt ctttttctca
     1921 atgtatgaga ggcgcattgg agttctgctg ttgatctcat taacacagac ttgcaggaag
     1981 cggcggcgga agtcaggcat acgctggtaa ctttgaggca gctggtaacg ctctatgatc
     2041 cagtcgattt tcagagacac gatgcctgag ccatccggct tacgatactg acacagggat
     2101 tcgtataaac gcatggcata cggattggtg atttcttttg tttcactaag ccgaaactgc
     2161 gtaaaccggt tctgtaaccc gataaagaag ggaatgagat atgggttgat atgtacactg
     2221 taaagccctc tggatggact gtgcgcacgt ttgataaacc aaggaaaaga ttcatagcct
     2281 ttttcatcac cggcatcctc ttcagggcga taaaaaacca cttccttccc cgcgaaactc
     2341 ttcaatgcct gccgtatatc cttactggct tccgcagagg tcaatccgaa tatttcagca
     2401 tatttagcaa catggatctc gcagataccg tcatgttcct gtagggtgcc atcagatttt
     2461 ctgatctggt caacgaacag atacagcata cgtttttgat cccgggagag actatatgcc
     2521 gcctcagtga ggtcgtttga ctggacgatt cgcgggctat ttttacgttt cttgtgattg
     2581 ataaccgctg tttccgccat gacagatcca tgtgaagtgt gacaagtttt tagattgtca
     2641 cactaaataa aaaagagtca ataagcaggg ataactttgt gaaaaaacag cttcttctga
     2701 gggcaatttg tcacagggtt aagggcaatt tgtcacagac aggactgtca tttgagggtg
     2761 atttgtcaca ctgaaagggc aatttgtcac aacaccttct gtagaaccag catggataaa
     2821 ggcctacaag gcgctctaaa aaagaagatc taaaaactat aaaaaaaata attataaaaa
     2881 tatccccgtg gataagtgga taaccccaag ggaagttttt tcaggcatcg tgtgtaagca
     2941 gaatatataa gtgctgttcc ctggtgcttc ctcgctcact cgaaattccc ggggatagct
     3001 ttatgcttgt aaaccgtttt gtgaaaaaat ttttaaaata aaaaagggga cctctagggt
     3061 ccccaattaa ttagtaatat aatctattaa aggtcattca aaaggtcatc caccggatca
     3121 attcccctgc tcgcgcaggc tgggtgccaa gctctcgggt aacatcaagg cccgatcctt
     3181 ggagcccttg ccctcccgca cgatgatcgt gccgtgatcg aaatccagat ccttgacccg
     3241 cagttgcaaa ccctcactga tccgattcat taatgcagct ggcacgacag gtttcccgac
     3301 tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt agctcactca ttaggcaccc
     3361 caggctttac actttatgct tccggctcgt atgttgtgtg gaattgtgag cggataacaa
     3421 tttcacacag gaaacagcta tgaccatgat tacgccaagc gcggtacctg tcgcggcaac
     3481 gcgctaacag acgtagtaag aaccaccagc attgtaatgc tggctaaagt cactttcctg
     3541 agctgtataa cgatgagcga ttttactttt tctggctatg aattggcctg ctttgtaaca
     3601 cactccggtc tatcccgtag cgccgggcat atcctgtcgc aatgtgcaaa tctcgcggca
     3661 acaaccagtg aatacttcat tcacaagcct caccgcctga tcgcggcaga aactggttat
     3721 agccaatcaa ccgtcgttcg tgcattccgt gaagctgtaa acaaaggtat cctgtctgta
     3781 gagattgtta tcggcgatca ccgtgaacgt cgcgctaacc tgtaccggtt tacaccatcc
     3841 tttttggcct tcgcacaaca agccaaaaat gcgctgatag aaagcaaatt aaagatctct
     3901 tcagcggcaa ccaaggttaa agctgttctc gctaagacat tggctttatt taatttttta
     3961 tccacacccc catgtcaaaa tgataccccc tccccctgtc aggatgacgt ggcaataaag
     4021 aataagaagt cacaagttaa aaaaacaaaa agatcagttt ccggcggtgc cggaacaacc
     4081 agcctcaaaa aattgacttc atggatcgct aaggcaaaag caaaggctga caatctgcgg
     4141 ttatccaaaa aacgcactca aaaacatgag ttcaagcaga aagtagaggc ggctgcgcgg
     4201 aaatatgctt acctgaagaa caagcgttcg cctgatattg gcgggatatc aaacttcgat
     4261 aacctaccgc attgcatgac ggtaaacgaa gctcttaatg cggttttagc caaaaataaa
     4321 gataacgaac aatggggtat accggcagga ttcagagggt aatgaattgc tctaattata
     4381 accatgcata ctttcaacac ctctagtttg ccatgaggca aactcatagg tgtcctggta
     4441 agaggacact gttgccaaaa ctggacgccc cattattgca attaataaac aactaacgga
     4501 caattctacc taacaataag tggagttgcg gccggccgcg atccgctagc aaagccacgt
     4561 tgtgtctcaa aatctctgat gttacattgc acaagataaa aatatatcat catgaacaat
     4621 aaaactgtct gcttacataa acagtaatac aaggggtgtt atgagccata ttcaacggga
     4681 aacgtcttgc tcgcgcccgc gattaaattc caacatggat gctgatttat atgggtataa
     4741 atgggctcgc gataatgtcg ggcaatcagg tgcgacaatc tatcgattgt atgggaagcc
     4801 cgatgcgcca gagttgtttc tgaaacatgg caaaggtagc gttgccaatg atgttacaga
     4861 tgagatggtc agactaaact ggctgacgga atttatgcct cttccgacca tcaagcattt
     4921 tatccgtact cctgatgatg catggttact caccactgcg atccccggga aaacagcatt
     4981 ccaggtatta gaagaatatc ctgattcagg tgaaaatatt gttgatgcgc tggcagtgtt
     5041 cctgcgccgg ttgcattcga ttcctgtttg taattgtcct tttaacagcg atcgcgtatt
     5101 tcgcctcgct caggcgcaat cacgaatgaa taacggtttg gttgatgcga gtgattttga
     5161 tgacgagcgt aatggctggc ctgttgaaca agtctggaaa gaaatgcata aactgttgcc
     5221 attctcaccg gattcagtcg tcactcatgg tgatttctca cttgataacc ttatttttga
     5281 cgaggggaaa ttaataggtt gtattgatgt tggacgagtc ggaatcgcag accgatacca
     5341 ggatcttgcc atcctatgga actgcctcgg tgagttttct ccttcattac agaaacggct
     5401 ttttcaaaaa tatggtattg ataatcctga tatgaataaa ttgcagtttc atttgatgct
     5461 cgatgagttt ttctaatcag aattggttaa ttggttgtaa cactggcgag ctctgccacc
     5521 tgacgtctaa gaaaccatta ttatcatgac attaacctat aaaaataggc gtatcacgag
     5581 gcagaatttc agataaaaaa aatccttagc tttcgctaag gatgatttct gcacctgcac
     5641 cagtcagtaa aacgacggcc agtagtcaaa agcctccgac cggaggcttt tgacttggtt
     5701 caggtggagt gggag
//
