LOCUS       Exported                6300 bp DNA     circular SYN 02-JUL-2021
DEFINITION  BBF10K_000533 in pOpen_v4.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6300)
  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..6300
                     /organism="BBF10K_000533 in pOpen_v4"
     primer_bind     21..51
                     /label=Gibson-for
     regulatory      46..55
                     /regulatory_class="other"
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             52..1806
                     /codon_start=1
                     /product="TACTILE T cell activation, increased late 
                     expression"
                     /label=CD96
                     /translation="MEKKWKYCAVYYIIQIHFVKGVWEKTVNTEENVYATLGSDVNLTC
                     QTQTVGFFVQMQWSKVTNKIDLIAVYHPQYGFYCAYGRPCESLVTFTETPENGSKWTLH
                     LRNMSCSVSGRYECMLVLYPEGIQTKIYNLLIQTHVTADEWNSNHTIEIEINQTLEIPC
                     FQNSSSKISSEFTYAWSVENSSTDSWVLLSKGIKEDNGTQETLISQNHLISNSTLLKDR
                     VKLGTDYRLHLSPVQIFDDGRKFSCHIRVGPNKILRSSTTVKVFAKPEIPVIVENNSTD
                     VLVERRFTCLLKNVFPKANITWFIDGSFLHDEKEGIYITNEERKGKDGFLELKSVLTRV
                     HSNKPAQSDNLTIWCMALSPVPGNKVWNISSEKITFLLGSEISSTDPPLSVTESTLDTQ
                     PSPASSVSPARYPATSSVTLVDVSALRPNTTPQPSNSSMTTRGFNYPWTSSGTDTKKSV
                     SRIPSETYSSSPSGAGSTLHDNVFTSTARAFSEVPTTANGSTKTNHVHITGIVVNKPKD
                     GMSWPVIVAALLFCCMILFGLGVRKWCQYQKEIMERPPPFKPPPPPIKYTCIQEPNESD
                     LPYHEMETL"
     primer_bind     complement(1807..1831)
                     /label=Gibson-rev
     terminator      1950..1997
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(2002..2018)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    2177..2427
                     /gene="incC"
                     /label=incC
                     /note="incompatibility region of the bacterial F plasmid"
     CDS             complement(2430..3185)
                     /codon_start=1
                     /gene="repE"
                     /product="replication initiation protein for the bacterial 
                     F plasmid"
                     /label=repE
                     /translation="MAETAVINHKKRKNSPRIVQSNDLTEAAYSLSRDQKRMLYLFVDQ
                     IRKSDGTLQEHDGICEIHVAKYAEIFGLTSAEASKDIRQALKSFAGKEVVFYRPEEDAG
                     DEKGYESFPWFIKRAHSPSRGLYSVHINPYLIPFFIGLQNRFTQFRLSETKEITNPYAM
                     RLYESLCQYRKPDGSGIVSLKIDWIIERYQLPQSYQRMPDFRRRFLQVCVNEINSRTPM
                     RLSYIEKKKGRQTTHIVFSFRDITSMTTG"
     rep_origin      3276..3495
                     /label=ori2
                     /note="secondary origin of replication for the bacterial F 
                     plasmid; also known as oriS"
     protein_bind    3915..3936
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        3951..3981
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3989..4005
                     /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     4013..4029
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             4102..4947
                     /codon_start=1
                     /label=repL
                     /translation="MLAKVTFLSCITMSDFTFSGYELACFVTHSGLSRSAGHILSQCAN
                     LAATTSEYFIHKPHRLIAAETGYSQSTVVRAFREAVNKGILSVEIVIGDHRERRANLYR
                     FTPSFLAFAQQAKNALIESKLKISSAATKVKAVLAKTLALFNFLSTPPCQNDTPSPCQD
                     DVAIKNKKSQVKKTKRSVSGGAGTTSLKKLTSWIAKAKAKADNLRLSKKRTQKHEFKQK
                     VEAAARKYAYLKNKRSPDIGGISNFDNLPHCMTVNEALNAVLAKNKDNEQWGIPAGFRG
                     "
     misc_feature    5055..5206
                     /label=nnpp promoters
     CDS             5246..6061
                     /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(6171..6214)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     primer_bind     6232..6248
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     terminator      6249..6280
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
ORIGIN
        1 aagaattact gacccctcgg cagactgagt cgcccggggg gatccgccac catggagaag
       61 aaatggaaat attgtgctgt ctactatatt attcaaattc attttgtcaa aggggtttgg
      121 gagaaaacag ttaatacaga agaaaatgtc tatgcaacgc tcggttcaga tgttaatctt
      181 acttgtcaaa cgcaaactgt tgggttcttt gttcaaatgc aatggtccaa agttacgaat
      241 aaaattgatc tcattgcagt ctatcatcca caatatggct tttattgtgc atatggaagg
      301 ccatgtgaat ccttggtcac gtttacagaa acacctgaaa atggttctaa atggacgctt
      361 catcttcgga atatgtcctg tagtgtctca ggacgctatg aatgtatgct cgtcctttat
      421 ccagaaggta ttcaaacaaa gatatataat ctcctgattc aaacgcatgt cacagctgat
      481 gaatggaatt ctaaccatac tatagaaata gaaataaatc agacgctcga aattccttgt
      541 tttcaaaatt ccagctccaa aattagtagt gaatttactt atgcatggtc cgttgagaat
      601 tcatccacag atagttgggt acttcttagt aaagggatta aagaagataa tgggacacaa
      661 gaaacactca tatctcaaaa tcatcttata tccaattcca cgcttctcaa agatcgcgtt
      721 aaattgggta cggattatcg ccttcatctc tcccctgtcc aaatctttga tgatggtcgg
      781 aaattttcct gtcatattag ggttggacct aataaaattc ttaggagttc cacgacggtc
      841 aaagttttcg ctaaaccaga aattcctgtt attgtcgaaa ataattccac ggatgttctt
      901 gttgagcgcc ggtttacttg tttgttgaag aatgtatttc ctaaagcaaa tataacttgg
      961 tttattgacg gatcattcct tcatgatgaa aaggaaggta tatatattac gaatgaagag
     1021 cgaaaaggaa aggatggatt tcttgaactt aaatctgtcc ttacacgcgt acatagcaat
     1081 aaacccgctc aatccgataa tttgacaatt tggtgtatgg cgttgtctcc tgtaccaggg
     1141 aataaagttt ggaatatttc atccgagaaa attacttttc tccttggatc agaaataagt
     1201 tcaacagatc caccactttc cgtcacagaa tctacactcg atacacaacc ttccccagca
     1261 tcttctgtaa gtccagctcg ctatcctgcg actagttcag tcactcttgt tgatgtaagt
     1321 gctttgcgac ctaatactac tcctcaaccg agtaattcta gtatgacaac acgcggattt
     1381 aattatcctt ggacttcttc tggaacagat acgaagaaat ctgtcagtcg cattccgtcc
     1441 gaaacttatt caagttcccc atccggtgca gggtctactc ttcatgataa tgtctttact
     1501 tctacagcac gagcattttc tgaagtacct actacagcaa atggttccac taaaacaaat
     1561 catgtccata ttacgggtat agtcgtcaat aaaccaaaag atggtatgtc atggccagtc
     1621 attgtcgcag ctctcttgtt ttgttgtatg attctctttg gactcggggt acggaaatgg
     1681 tgtcaatatc aaaaggaaat tatggaaagg ccaccacctt ttaaaccacc accaccacca
     1741 attaaatata cttgtattca agaacctaat gaaagtgatc ttccgtatca cgagatggag
     1801 accctgggat ccgccaccaa cttcagcctg ctagtattgc cggactaaac ccagcctcaa
     1861 caaacctaaa cgttagggat aacagggtaa ttgacatatc tccggcctga cacgctgagg
     1921 tgtcaatcgt cggagccgct gagcaataac tagcataacc ccttggggcc tctaaacggg
     1981 tcttgagggg ttttttgcat ggtcatagct gtttcctgag agcttggcag gtgtcgcgta
     2041 tgcatcgatc acgtgtccac gttctttaat agtggactct tgttccaaac tggaacaaca
     2101 ctcggatcga tccggcgcgc accgtgggaa aaactccagg tagaggtaca cacgcggata
     2161 gccaattcag agtaataaac tgtgataatc aaccctcatc aatgatgacg aactaacccc
     2221 cgatatcagg tcacatgacg aagggaaaga gaaggaaatc aactgtgaca aactgccctc
     2281 aaatttggct tccttaaaaa ttacagttca aaaagtatga gaaaatccat gcaggctgaa
     2341 ggaaacagca aaactgtgac aaattaccct cagtaggtca gaacaaatgt gacgaaccac
     2401 cctcaaatct gtgacagata accctcagac tatcctgtcg tcatggaagt gatatcgcgg
     2461 aaggaaaata cgatatgagt cgtctggcgg cctttctttt tctcaatgta tgagaggcgc
     2521 attggagttc tgctgttgat ctcattaaca cagacttgca ggaagcggcg gcggaagtca
     2581 ggcatacgct ggtaactttg aggcagctgg taacgctcta tgatccagtc gattttcaga
     2641 gacacgatgc ctgagccatc cggcttacga tactgacaca gggattcgta taaacgcatg
     2701 gcatacggat tggtgatttc ttttgtttca ctaagccgaa actgcgtaaa ccggttctgt
     2761 aacccgataa agaagggaat gagatatggg ttgatatgta cactgtaaag ccctctggat
     2821 ggactgtgcg cacgtttgat aaaccaagga aaagattcat agcctttttc atcaccggca
     2881 tcctcttcag ggcgataaaa aaccacttcc ttccccgcga aactcttcaa tgcctgccgt
     2941 atatccttac tggcttccgc agaggtcaat ccgaatattt cagcatattt agcaacatgg
     3001 atctcgcaga taccgtcatg ttcctgtagg gtgccatcag attttctgat ctggtcaacg
     3061 aacagataca gcatacgttt ttgatcccgg gagagactat atgccgcctc agtgaggtcg
     3121 tttgactgga cgattcgcgg gctattttta cgtttcttgt gattgataac cgctgtttcc
     3181 gccatgacag atccatgtga agtgtgacaa gtttttagat tgtcacacta aataaaaaag
     3241 agtcaataag cagggataac tttgtgaaaa aacagcttct tctgagggca atttgtcaca
     3301 gggttaaggg caatttgtca cagacaggac tgtcatttga gggtgatttg tcacactgaa
     3361 agggcaattt gtcacaacac cttctgtaga accagcatgg ataaaggcct acaaggcgct
     3421 ctaaaaaaga agatctaaaa actataaaaa aaataattat aaaaatatcc ccgtggataa
     3481 gtggataacc ccaagggaag ttttttcagg catcgtgtgt aagcagaata tataagtgct
     3541 gttccctggt gcttcctcgc tcactcgaaa ttcccgggga tagctttatg cttgtaaacc
     3601 gttttgtgaa aaaattttta aaataaaaaa ggggacctct agggtcccca attaattagt
     3661 aatataatct attaaaggtc attcaaaagg tcatccaccg gatcaattcc cctgctcgcg
     3721 caggctgggt gccaagctct cgggtaacat caaggcccga tccttggagc ccttgccctc
     3781 ccgcacgatg atcgtgccgt gatcgaaatc cagatccttg acccgcagtt gcaaaccctc
     3841 actgatccga ttcattaatg cagctggcac gacaggtttc ccgactggaa agcgggcagt
     3901 gagcgcaacg caattaatgt gagttagctc actcattagg caccccaggc tttacacttt
     3961 atgcttccgg ctcgtatgtt gtgtggaatt gtgagcggat aacaatttca cacaggaaac
     4021 agctatgacc atgattacgc caagcgcggt acctgtcgcg gcaacgcgct aacagacgta
     4081 gtaagaacca ccagcattgt aatgctggct aaagtcactt tcctgagctg tataacgatg
     4141 agcgatttta ctttttctgg ctatgaattg gcctgctttg taacacactc cggtctatcc
     4201 cgtagcgccg ggcatatcct gtcgcaatgt gcaaatctcg cggcaacaac cagtgaatac
     4261 ttcattcaca agcctcaccg cctgatcgcg gcagaaactg gttatagcca atcaaccgtc
     4321 gttcgtgcat tccgtgaagc tgtaaacaaa ggtatcctgt ctgtagagat tgttatcggc
     4381 gatcaccgtg aacgtcgcgc taacctgtac cggtttacac catccttttt ggccttcgca
     4441 caacaagcca aaaatgcgct gatagaaagc aaattaaaga tctcttcagc ggcaaccaag
     4501 gttaaagctg ttctcgctaa gacattggct ttatttaatt ttttatccac acccccatgt
     4561 caaaatgata ccccctcccc ctgtcaggat gacgtggcaa taaagaataa gaagtcacaa
     4621 gttaaaaaaa caaaaagatc agtttccggc ggtgccggaa caaccagcct caaaaaattg
     4681 acttcatgga tcgctaaggc aaaagcaaag gctgacaatc tgcggttatc caaaaaacgc
     4741 actcaaaaac atgagttcaa gcagaaagta gaggcggctg cgcggaaata tgcttacctg
     4801 aagaacaagc gttcgcctga tattggcggg atatcaaact tcgataacct accgcattgc
     4861 atgacggtaa acgaagctct taatgcggtt ttagccaaaa ataaagataa cgaacaatgg
     4921 ggtataccgg caggattcag agggtaatga attgctctaa ttataaccat gcatactttc
     4981 aacacctcta gtttgccatg aggcaaactc ataggtgtcc tggtaagagg acactgttgc
     5041 caaaactgga cgccccatta ttgcaattaa taaacaacta acggacaatt ctacctaaca
     5101 ataagtggag ttgcggccgg ccgcgatccg ctagcaaagc cacgttgtgt ctcaaaatct
     5161 ctgatgttac attgcacaag ataaaaatat atcatcatga acaataaaac tgtctgctta
     5221 cataaacagt aatacaaggg gtgttatgag ccatattcaa cgggaaacgt cttgctcgcg
     5281 cccgcgatta aattccaaca tggatgctga tttatatggg tataaatggg ctcgcgataa
     5341 tgtcgggcaa tcaggtgcga caatctatcg attgtatggg aagcccgatg cgccagagtt
     5401 gtttctgaaa catggcaaag gtagcgttgc caatgatgtt acagatgaga tggtcagact
     5461 aaactggctg acggaattta tgcctcttcc gaccatcaag cattttatcc gtactcctga
     5521 tgatgcatgg ttactcacca ctgcgatccc cgggaaaaca gcattccagg tattagaaga
     5581 atatcctgat tcaggtgaaa atattgttga tgcgctggca gtgttcctgc gccggttgca
     5641 ttcgattcct gtttgtaatt gtccttttaa cagcgatcgc gtatttcgcc tcgctcaggc
     5701 gcaatcacga atgaataacg gtttggttga tgcgagtgat tttgatgacg agcgtaatgg
     5761 ctggcctgtt gaacaagtct ggaaagaaat gcataaactg ttgccattct caccggattc
     5821 agtcgtcact catggtgatt tctcacttga taaccttatt tttgacgagg ggaaattaat
     5881 aggttgtatt gatgttggac gagtcggaat cgcagaccga taccaggatc ttgccatcct
     5941 atggaactgc ctcggtgagt tttctccttc attacagaaa cggctttttc aaaaatatgg
     6001 tattgataat cctgatatga ataaattgca gtttcatttg atgctcgatg agtttttcta
     6061 atcagaattg gttaattggt tgtaacactg gcgagctctg ccacctgacg tctaagaaac
     6121 cattattatc atgacattaa cctataaaaa taggcgtatc acgaggcaga atttcagata
     6181 aaaaaaatcc ttagctttcg ctaaggatga tttctgcacc tgcaccagtc agtaaaacga
     6241 cggccagtag tcaaaagcct ccgaccggag gcttttgact tggttcaggt ggagtgggag
//
