LOCUS       Exported                5640 bp DNA     circular SYN 02-JUL-2021
DEFINITION  BBF10K_000522 in pOpen_v4.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5640)
  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..5640
                     /organism="BBF10K_000522 in pOpen_v4"
     primer_bind     21..51
                     /label=Gibson-for
     CDS             52..1146
                     /codon_start=1
                     /product="Homo sapiens CD244 molecule"
                     /label=2B4
                     /translation="MLGQVVTLILLLLLKVYQGKGCQGSADHVVSISGVPLQLQPNSIQ
                     TKVDSIAWKKLLPSQNGFHHILKWENGSLPSNTSNDRFSFIVKNLSLLIKAAQQQDSGL
                     YCLEVTSISGKVQTATFQVFVFDKVEKPRLQGQGKILDRGRCQVALSCLVSRDGNVSYA
                     WYRGSKLIQTAGNLTYLDEEVDINGTHTYTCNVSNPVSWESHTLNLTQDCQNAHQEFRF
                     WPFLVIIVILSALFLGTLACFCVWRRKRKEKQSETSPKEFLTIYEDVKDLKTRRNHEQE
                     QTFPGGGSTIYSMIQSQSSAPTSQEPAYTLYSLIQPSRKSGSRKRNHSPSFNSTIYEVI
                     GKSQPKAQNPARLSRKELENFDVYS"
     primer_bind     complement(1147..1171)
                     /label=Gibson-rev
     terminator      1290..1337
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(1342..1358)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    1517..1767
                     /gene="incC"
                     /label=incC
                     /note="incompatibility region of the bacterial F plasmid"
     CDS             complement(1770..2525)
                     /codon_start=1
                     /gene="repE"
                     /product="replication initiation protein for the bacterial 
                     F plasmid"
                     /label=repE
                     /translation="MAETAVINHKKRKNSPRIVQSNDLTEAAYSLSRDQKRMLYLFVDQ
                     IRKSDGTLQEHDGICEIHVAKYAEIFGLTSAEASKDIRQALKSFAGKEVVFYRPEEDAG
                     DEKGYESFPWFIKRAHSPSRGLYSVHINPYLIPFFIGLQNRFTQFRLSETKEITNPYAM
                     RLYESLCQYRKPDGSGIVSLKIDWIIERYQLPQSYQRMPDFRRRFLQVCVNEINSRTPM
                     RLSYIEKKKGRQTTHIVFSFRDITSMTTG"
     rep_origin      2616..2835
                     /label=ori2
                     /note="secondary origin of replication for the bacterial F 
                     plasmid; also known as oriS"
     protein_bind    3255..3276
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        3291..3321
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3329..3345
                     /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     3353..3369
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             3442..4287
                     /codon_start=1
                     /label=repL
                     /translation="MLAKVTFLSCITMSDFTFSGYELACFVTHSGLSRSAGHILSQCAN
                     LAATTSEYFIHKPHRLIAAETGYSQSTVVRAFREAVNKGILSVEIVIGDHRERRANLYR
                     FTPSFLAFAQQAKNALIESKLKISSAATKVKAVLAKTLALFNFLSTPPCQNDTPSPCQD
                     DVAIKNKKSQVKKTKRSVSGGAGTTSLKKLTSWIAKAKAKADNLRLSKKRTQKHEFKQK
                     VEAAARKYAYLKNKRSPDIGGISNFDNLPHCMTVNEALNAVLAKNKDNEQWGIPAGFRG
                     "
     misc_feature    4395..4546
                     /label=nnpp promoters
     CDS             4586..5401
                     /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(5511..5554)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     primer_bind     5572..5588
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     terminator      5589..5620
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
ORIGIN
        1 aagaattact gacccctcgg cagactgagt cgcccggggg gatccgccac catgctgggc
       61 caggtggtga ccctgatcct gctgctgctg ctgaaggtgt accagggcaa gggctgccag
      121 ggcagcgccg accacgtggt gagcatcagc ggcgtgcccc tgcagctgca gcccaacagc
      181 atccagacca aggtggacag catcgcctgg aagaagctgc tgcccagcca gaacggcttc
      241 caccacatcc tgaagtggga gaacggcagc ctgcccagca acaccagcaa cgacagattc
      301 agcttcatcg tgaagaacct gagcctgctg atcaaggccg cccagcagca ggacagcggc
      361 ctgtactgcc tggaggtgac cagcatcagc ggcaaggtgc agaccgccac cttccaggtg
      421 ttcgtgttcg acaaggtgga gaagcccaga ctgcagggcc agggcaagat cctggacaga
      481 ggcagatgcc aggtggccct gagctgcctg gtgagcagag acggcaacgt gagctacgcc
      541 tggtacagag gcagcaagct gatccagacc gccggcaacc tgacctacct ggacgaggag
      601 gtggacatca acggcaccca cacctacacc tgcaacgtga gcaaccccgt gagctgggag
      661 agccacaccc tgaacctgac ccaggactgc cagaacgccc accaggagtt cagattctgg
      721 cccttcctgg tgatcatcgt gatcctgagc gccctgttcc tgggcaccct ggcctgcttc
      781 tgcgtgtgga gaagaaagag aaaggagaag cagagcgaga ccagccccaa ggagttcctg
      841 accatctacg aggacgtgaa ggacctgaag accagaagaa accacgagca ggagcagacc
      901 ttccccggcg gcggcagcac catctacagc atgatccaga gccagagcag cgcccccacc
      961 agccaggagc ccgcctacac cctgtacagc ctgatccagc ccagcagaaa gagcggcagc
     1021 agaaagagaa accacagccc cagcttcaac agcaccatct acgaggtgat cggcaagagc
     1081 cagcccaagg cccagaaccc cgccagactg agcagaaagg agctggagaa cttcgacgtg
     1141 tacagcggat ccgccaccaa cttcagcctg ctagctataa gaattgccgg gcctctgggc
     1201 gttcctcgcg ttatagggat aacagggtaa tctgatcgga cgctagaact ggcgctgagg
     1261 tgtcaatcgt cggagccgct gagcaataac tagcataacc ccttggggcc tctaaacggg
     1321 tcttgagggg ttttttgcat ggtcatagct gtttcctgag agcttggcag gtgtcgcgta
     1381 tgcatcgatc acgtgtccac gttctttaat agtggactct tgttccaaac tggaacaaca
     1441 ctcggatcga tccggcgcgc accgtgggaa aaactccagg tagaggtaca cacgcggata
     1501 gccaattcag agtaataaac tgtgataatc aaccctcatc aatgatgacg aactaacccc
     1561 cgatatcagg tcacatgacg aagggaaaga gaaggaaatc aactgtgaca aactgccctc
     1621 aaatttggct tccttaaaaa ttacagttca aaaagtatga gaaaatccat gcaggctgaa
     1681 ggaaacagca aaactgtgac aaattaccct cagtaggtca gaacaaatgt gacgaaccac
     1741 cctcaaatct gtgacagata accctcagac tatcctgtcg tcatggaagt gatatcgcgg
     1801 aaggaaaata cgatatgagt cgtctggcgg cctttctttt tctcaatgta tgagaggcgc
     1861 attggagttc tgctgttgat ctcattaaca cagacttgca ggaagcggcg gcggaagtca
     1921 ggcatacgct ggtaactttg aggcagctgg taacgctcta tgatccagtc gattttcaga
     1981 gacacgatgc ctgagccatc cggcttacga tactgacaca gggattcgta taaacgcatg
     2041 gcatacggat tggtgatttc ttttgtttca ctaagccgaa actgcgtaaa ccggttctgt
     2101 aacccgataa agaagggaat gagatatggg ttgatatgta cactgtaaag ccctctggat
     2161 ggactgtgcg cacgtttgat aaaccaagga aaagattcat agcctttttc atcaccggca
     2221 tcctcttcag ggcgataaaa aaccacttcc ttccccgcga aactcttcaa tgcctgccgt
     2281 atatccttac tggcttccgc agaggtcaat ccgaatattt cagcatattt agcaacatgg
     2341 atctcgcaga taccgtcatg ttcctgtagg gtgccatcag attttctgat ctggtcaacg
     2401 aacagataca gcatacgttt ttgatcccgg gagagactat atgccgcctc agtgaggtcg
     2461 tttgactgga cgattcgcgg gctattttta cgtttcttgt gattgataac cgctgtttcc
     2521 gccatgacag atccatgtga agtgtgacaa gtttttagat tgtcacacta aataaaaaag
     2581 agtcaataag cagggataac tttgtgaaaa aacagcttct tctgagggca atttgtcaca
     2641 gggttaaggg caatttgtca cagacaggac tgtcatttga gggtgatttg tcacactgaa
     2701 agggcaattt gtcacaacac cttctgtaga accagcatgg ataaaggcct acaaggcgct
     2761 ctaaaaaaga agatctaaaa actataaaaa aaataattat aaaaatatcc ccgtggataa
     2821 gtggataacc ccaagggaag ttttttcagg catcgtgtgt aagcagaata tataagtgct
     2881 gttccctggt gcttcctcgc tcactcgaaa ttcccgggga tagctttatg cttgtaaacc
     2941 gttttgtgaa aaaattttta aaataaaaaa ggggacctct agggtcccca attaattagt
     3001 aatataatct attaaaggtc attcaaaagg tcatccaccg gatcaattcc cctgctcgcg
     3061 caggctgggt gccaagctct cgggtaacat caaggcccga tccttggagc ccttgccctc
     3121 ccgcacgatg atcgtgccgt gatcgaaatc cagatccttg acccgcagtt gcaaaccctc
     3181 actgatccga ttcattaatg cagctggcac gacaggtttc ccgactggaa agcgggcagt
     3241 gagcgcaacg caattaatgt gagttagctc actcattagg caccccaggc tttacacttt
     3301 atgcttccgg ctcgtatgtt gtgtggaatt gtgagcggat aacaatttca cacaggaaac
     3361 agctatgacc atgattacgc caagcgcggt acctgtcgcg gcaacgcgct aacagacgta
     3421 gtaagaacca ccagcattgt aatgctggct aaagtcactt tcctgagctg tataacgatg
     3481 agcgatttta ctttttctgg ctatgaattg gcctgctttg taacacactc cggtctatcc
     3541 cgtagcgccg ggcatatcct gtcgcaatgt gcaaatctcg cggcaacaac cagtgaatac
     3601 ttcattcaca agcctcaccg cctgatcgcg gcagaaactg gttatagcca atcaaccgtc
     3661 gttcgtgcat tccgtgaagc tgtaaacaaa ggtatcctgt ctgtagagat tgttatcggc
     3721 gatcaccgtg aacgtcgcgc taacctgtac cggtttacac catccttttt ggccttcgca
     3781 caacaagcca aaaatgcgct gatagaaagc aaattaaaga tctcttcagc ggcaaccaag
     3841 gttaaagctg ttctcgctaa gacattggct ttatttaatt ttttatccac acccccatgt
     3901 caaaatgata ccccctcccc ctgtcaggat gacgtggcaa taaagaataa gaagtcacaa
     3961 gttaaaaaaa caaaaagatc agtttccggc ggtgccggaa caaccagcct caaaaaattg
     4021 acttcatgga tcgctaaggc aaaagcaaag gctgacaatc tgcggttatc caaaaaacgc
     4081 actcaaaaac atgagttcaa gcagaaagta gaggcggctg cgcggaaata tgcttacctg
     4141 aagaacaagc gttcgcctga tattggcggg atatcaaact tcgataacct accgcattgc
     4201 atgacggtaa acgaagctct taatgcggtt ttagccaaaa ataaagataa cgaacaatgg
     4261 ggtataccgg caggattcag agggtaatga attgctctaa ttataaccat gcatactttc
     4321 aacacctcta gtttgccatg aggcaaactc ataggtgtcc tggtaagagg acactgttgc
     4381 caaaactgga cgccccatta ttgcaattaa taaacaacta acggacaatt ctacctaaca
     4441 ataagtggag ttgcggccgg ccgcgatccg ctagcaaagc cacgttgtgt ctcaaaatct
     4501 ctgatgttac attgcacaag ataaaaatat atcatcatga acaataaaac tgtctgctta
     4561 cataaacagt aatacaaggg gtgttatgag ccatattcaa cgggaaacgt cttgctcgcg
     4621 cccgcgatta aattccaaca tggatgctga tttatatggg tataaatggg ctcgcgataa
     4681 tgtcgggcaa tcaggtgcga caatctatcg attgtatggg aagcccgatg cgccagagtt
     4741 gtttctgaaa catggcaaag gtagcgttgc caatgatgtt acagatgaga tggtcagact
     4801 aaactggctg acggaattta tgcctcttcc gaccatcaag cattttatcc gtactcctga
     4861 tgatgcatgg ttactcacca ctgcgatccc cgggaaaaca gcattccagg tattagaaga
     4921 atatcctgat tcaggtgaaa atattgttga tgcgctggca gtgttcctgc gccggttgca
     4981 ttcgattcct gtttgtaatt gtccttttaa cagcgatcgc gtatttcgcc tcgctcaggc
     5041 gcaatcacga atgaataacg gtttggttga tgcgagtgat tttgatgacg agcgtaatgg
     5101 ctggcctgtt gaacaagtct ggaaagaaat gcataaactg ttgccattct caccggattc
     5161 agtcgtcact catggtgatt tctcacttga taaccttatt tttgacgagg ggaaattaat
     5221 aggttgtatt gatgttggac gagtcggaat cgcagaccga taccaggatc ttgccatcct
     5281 atggaactgc ctcggtgagt tttctccttc attacagaaa cggctttttc aaaaatatgg
     5341 tattgataat cctgatatga ataaattgca gtttcatttg atgctcgatg agtttttcta
     5401 atcagaattg gttaattggt tgtaacactg gcgagctctg ccacctgacg tctaagaaac
     5461 cattattatc atgacattaa cctataaaaa taggcgtatc acgaggcaga atttcagata
     5521 aaaaaaatcc ttagctttcg ctaaggatga tttctgcacc tgcaccagtc agtaaaacga
     5581 cggccagtag tcaaaagcct ccgaccggag gcttttgact tggttcaggt ggagtgggag
//
