LOCUS       Exported                4985 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Express soluble His-tagged zebrafish erythropoietin in E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    pHis-zfEpoa
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4985)
  AUTHORS   Stachura DL, Reyes JR, Bartunek P, Paw BH, Zon LI, Traver D
  TITLE     Zebrafish kidney stromal cell lines support multilineage 
            hematopoiesis.
  JOURNAL   Blood. 2009 Jul 9;114(2):279-89. doi: 10.1182/blood-2009-02-203638. 
            Epub 2009 May 11.
  PUBMED    19433857
REFERENCE   2  (bases 1 to 4985)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..4985
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     promoter        27..131
                     /gene="bla"
                     /label=AmpR promoter
     CDS             132..992
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(350..369)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1163..1751
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1652..1671
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1905..1922
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    1937..2076
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2002..2021)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2162..2184
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2178..2369)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     3827..3846
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     3930..3949
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3930..3948
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             3980..4002
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             4019..4036
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     promoter        complement(4871..4899)
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(4926..4944)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
       61 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
      121 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat
      181 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc
      241 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga
      301 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg
      361 cggtattatc ccgtgttgac gccgggcaag agcaactcgg tcgccgcata cactattctc
      421 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag
      481 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc
      541 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg
      601 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg
      661 acaccacgat gcctgcagca atggcaacaa cgttgcgcaa actattaact ggcgaactac
      721 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac
      781 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg
      841 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg
      901 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg
      961 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac
     1021 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
     1081 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     1141 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     1201 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     1261 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc cttctagtgt
     1321 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     1381 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     1441 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     1501 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     1561 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     1621 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     1681 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     1741 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     1801 ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct
     1861 ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg
     1921 aggaagcgga agagcgcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac
     1981 accgcaatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagtatac
     2041 actccgctat cgctacgtga ctgggtcatg gctgcgcccc gacacccgcc aacacccgct
     2101 gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc
     2161 tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gaggcagctg
     2221 cggtaaagct catcagcgtg gtcgtgaagc gattcacaga tgtctgcctg ttcatccgcg
     2281 tccagctcgt tgagtttctc cagaagcgtt aatgtctggc ttctgataaa gcgggccatg
     2341 ttaagggcgg ttttttcctg tttggtcact gatgcctccg tgtaaggggg atttctgttc
     2401 atgggggtaa tgataccgat gaaacgagag aggatgctca cgatacgggt tactgatgat
     2461 gaacatgccc ggttactgga acgttgtgag ggtaaacaac tggcggtatg gatgcggcgg
     2521 gaccagagaa aaatcactca gggtcaatgc cagcgcttcg ttaatacaga tgtaggtgtt
     2581 ccacagggta gccagcagca tcctgcgatg cagatccgga acataatggt gcagggcgct
     2641 gacttccgcg tttccagact ttacgaaaca cggaaaccga agaccattca tgttgttgct
     2701 caggtcgcag acgttttgca gcagcagtcg cttcacgttc gctcgcgtat cggtgattca
     2761 ttctgctaac cagtaaggca accccgccag cctagccggg tcctcaacga caggagcacg
     2821 atcatgcgca cccgtggcca ggacccaacg ctgcccgaga tgcgccgcgt gcggctgctg
     2881 gagatggcgg acgcgatgga tatgttctgc caagggttgg tttgcgcatt cacagttctc
     2941 cgcaagaatt gattggctcc aattcttgga gtggtgaatc cgttagcgag gtgccgccgg
     3001 cttccattca ggtcgaggtg gcccggctcc atgcaccgcg acgcaacgcg gggaggcaga
     3061 caaggtatag ggcggcgcct acaatccatg ccaacccgtt ccatgtgctc gccgaggcgg
     3121 cataaatcgc cgtgacgatc agcggtccaa tgatcgaagt taggctggta agagccgcga
     3181 gcgatccttg aagctgtccc tgatggtcgt catctacctg cctggacagc atggcctgca
     3241 acgcgggcat cccgatgccg ccggaagcga gaagaatcat aatggggaag gccatccagc
     3301 ctcgcgtcgc gaacgccagc aagacgtagc ccagcgcgtc ggccgccatg ccggcgataa
     3361 tggcctgctt ctcgccgaaa cgtttggtgg cgggaccagt gacgaaggct tgagcgaggg
     3421 cgtgcaagat tccgaatacc gcaagcgaca ggccgatcat cgtcgcgctc cagcgaaagc
     3481 ggtcctcgcc gaaaatgacc cagagcgctg ccggcacctg tcctacgagt tgcatgataa
     3541 agaagacagt cataagtgcg gcgacgatag tcatgccccg cgcccaccgg aaggagctga
     3601 ctgggttgaa ggctctcaag ggcatcggtc gacgctctcc cttatgcgac tcctgcatta
     3661 ggaagcagcc cagtagtagg ttgaggccgt tgagcaccgc cgccgcaagg aatggtgcat
     3721 gcaaggagat ggcgcccaac agtcccccgg ccacggggcc tgccaccata cccacgccga
     3781 aacaagcgct catgagcccg aagtggcgag cccgatcttc cccatcggtg atgtcggcga
     3841 tataggcgcc agcaaccgca cctgtggcgc cggtgatgcc ggccacgatg cgtccggcgt
     3901 agaggatcga gatctcgatc ccgcgaaatt aatacgactc actataggga gaccacaacg
     3961 gtttccctct agaaataatt ttgtttaact ttaagaagga gatatacata tggctagcca
     4021 ccatcaccat caccatggtg gatcctcccc attacgcccc atctgtgacc tgcgcgtcct
     4081 cgaccatttc attaaggagg catgggatgc agaggctgct atgagaactt gtaaggacga
     4141 ttgcagcatt gcaacgaacg tcactgttcc tttgaccaga gtcgattttg aagtctggga
     4201 agcgatgaat atagaggagc aagctcagga ggtccagtca ggcttacaca tgctgaacga
     4261 ggccatcggc tcattacaga tatctaatca gactgaagtg cttcagtctc acatagatgc
     4321 cagtattaga aacatcgcca gcatcagaca agtgctgcga agtctcagca taccggaata
     4381 tgtacctcca accagtagtg gagaagacaa ggagacacag aaaatatcct cgatctcaga
     4441 gctgtttcag gtccatgtca actttcttcg gggaaaagcg cgtctgctgc tcgccaatgc
     4501 acctgtctgt cgacagggtg tcagctgagg atccacagga cgggtgtggt cgccatgatc
     4561 gcgtagtcga tagtggctcc aagtagcgaa gcgagcagga ctgggcggcg gccaaagcgg
     4621 tcggacagtg ctccgagaac gggtgcgcat agaaattgca tcaacgcata tagcgctagc
     4681 agcacgccat agtgactggc gatgctgtcg gaatggacga tatcccgcaa gaggcccggc
     4741 agtaccggca taaccaagcc tatgcctaca gcatccaggg tgacggtgcc gaggatgacg
     4801 atgagcgcat tgttagattt catacacggt gcctgactgc gttagcaatt taactgtgat
     4861 aaactaccgc attaaagctt atcgatgata agctgtcaaa catgagaatt cttgaagacg
     4921 aaagggcctc gtgatacgcc tatttttata ggttaatgtc atgataataa tggtttctta
     4981 gacgt
//
