LOCUS       pET-28a_PA1472_Nterm_Hi 5848 bp ds-DNA     circular     10-FEB-2023
DEFINITION  synthetic DNA
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
COMMENT     Imported using the Genbank importer. File name:
            pET-28a_Nhis_PA1472.txt NEBuilder-generated oligos (UPPERCASE =
            gene-specific, lowercase = overlap) PA1472_fwd:
            tggtgccgcgcggcagccatATGAACTTTGTTTTTCCGC PA1472_fwd 3'Tm: 58.3
            3'Ta: 59.3 PA1472_rev: tggtggtggtggtgctcgagTTAGCGATTACGCCAATG
            PA1472_rev 3'Tm: 59.5  3'Ta: 59.3 pET-28a_Nhis_fwd:
            CTCGAGCACCACCACCACCACC pET-28a_Nhis_fwd 3'Tm: 75.5  3'Ta: 72.0
            pET-28a_Nhis_rev: ATGGCTGCCGCGCGGCAC pET-28a_Nhis_rev 3'Tm: 79.0
            3'Ta: 72.0
FEATURES             Location/Qualifiers
     primer_bind     244..263
                     /label="T7"
                     /ApEinfo_revcolor="#85dae9"
                     /ApEinfo_fwdcolor="#85dae9"
     misc_binding    263..285
                     /label="LacO"
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
     protein_bind    267..283
                     /label="lac operator"
                     /ApEinfo_revcolor="#d59687"
                     /ApEinfo_fwdcolor="#d59687"
     primer_bind     complement(305..333)
                     /label="pET28a Cterm Rev"
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
     CDS             331..945
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
                     /translation="MGSSHHHHHHSSGLVPRGSHMNFVFPHPLDTPRLRLRTFREEDAAPLFAMMSDPEVMRYWNTPPWTTPAQAREAILRDSQALVDGEYLNLAIERREDGQLLGSCILFHFEKGSRRAELGYCLARAAQGRGYMGEALRRLLAFAFDEIDLNRLEAEIDPRNRPSAASLERLGFRQEGLLAQRWIVSGEVSDSALYGLLAEHWRNR*"
     primer_bind     371..409
                     /label="PA1472_fwd"
                     /ApEinfo_revcolor="#d6b295"
                     /ApEinfo_fwdcolor="#d6b295"
                     /note="gene-specific Tm: 58.3 Ta: 59.3"
                     /note="overlap: tggtgccgcgcggcagccat"
                     /note="gene-specific primer: ATGAACTTTGTTTTTCCGC"
     primer_bind     complement(373..390)
                     /label="pET-28 Rev"
                     /ApEinfo_revcolor="#75c6a9"
                     /ApEinfo_fwdcolor="#75c6a9"
     primer_bind     complement(373..390)
                     /label="pET-28a_Nhis_rev"
                     /ApEinfo_revcolor="#f8d3a9"
                     /ApEinfo_fwdcolor="#f8d3a9"
                     /note="gene-specific Tm: 79.0 Ta: 72.0"
                     /note="gene-specific primer: ATGGCTGCCGCGCGGCAC"
     gene            391..945
                     /label="PA1472"
                     /ApEinfo_revcolor="#faac61"
                     /ApEinfo_fwdcolor="#faac61"
     primer_bind     complement(928..965)
                     /label="PA1472_rev"
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
                     /note="gene-specific Tm: 59.5 Ta: 59.3"
                     /note="overlap: tggtggtggtggtgctcgag"
                     /note="gene-specific primer: TTAGCGATTACGCCAATG"
     gene            946..390
                     /label="pET-28a_Nhis"
                     /ApEinfo_revcolor="#9eafd2"
                     /ApEinfo_fwdcolor="#9eafd2"
     primer_bind     946..963
                     /label="pET-28a Cterm For"
                     /ApEinfo_revcolor="#85dae9"
                     /ApEinfo_fwdcolor="#85dae9"
     primer_bind     946..966
                     /label="pET-28a For"
                     /ApEinfo_revcolor="#ffef86"
                     /ApEinfo_fwdcolor="#ffef86"
     primer_bind     946..967
                     /label="pET-28a_Nhis_fwd"
                     /ApEinfo_revcolor="#faac61"
                     /ApEinfo_fwdcolor="#faac61"
                     /note="gene-specific Tm: 75.5 Ta: 72.0"
                     /note="gene-specific primer: CTCGAGCACCACCACCACCACC"
     CDS             952..969
                     /label="6xHis"
                     /ApEinfo_revcolor="#ff9ccd"
                     /ApEinfo_fwdcolor="#ff9ccd"
     CDS             952..969
                     /label="6x His Tag"
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
     rep_origin      1120..1575
                     /label="M13 origin"
                     /ApEinfo_revcolor="#9eafd2"
                     /ApEinfo_fwdcolor="#9eafd2"
     rep_origin      1137..1443
                     /label="F1 ori"
                     /ApEinfo_revcolor="#9eafd2"
                     /ApEinfo_fwdcolor="#9eafd2"
     CDS             complement(1671..2483)
                     /label="KanR"
                     /ApEinfo_revcolor="#ffef86"
                     /ApEinfo_fwdcolor="#ffef86"
     rep_origin      2529..3211
                     /label="ColE1 origin"
                     /ApEinfo_revcolor="#9eafd2"
                     /ApEinfo_fwdcolor="#9eafd2"
     primer_bind     3279..3299
                     /label="Seq Primer for Rev"
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
     primer_bind     complement(3378..3398)
                     /label="2,900 R"
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
ORIGIN
        1 ccgttgagca ccgccgccgc aaggaatggt gcatgcaagg agatggcgcc caacagtccc
       61 ccggccacgg ggcctgccac catacccacg ccgaaacaag cgctcatgag cccgaagtgg
      121 cgagcccgat cttccccatc ggtgatgtcg gcgatatagg cgccagcaac cgcacctgtg
      181 gcgccggtga tgccggccac gatgcgtccg gcgtagagga tcgagatctc gatcccgcga
      241 aattaatacg actcactata ggggaattgt gagcggataa caattcccct ctagaaataa
      301 ttttgtttaa ctttaagaag gagatatacc atgggcagca gccatcatca tcatcatcac
      361 agcagcggcc tggtgccgcg cggcagccat atgaactttg tttttccgca tccgctggat
      421 acaccgcgtc tgcgtctgcg cacctttcgt gaagaagatg cagcaccgct gtttgcaatg
      481 atgagcgatc cggaagttat gcgttattgg aatacccctc cgtggaccac accggcacag
      541 gcacgtgaag caattctgcg tgatagccag gcactggttg atggtgaata tctgaatctg
      601 gcaattgaac gtcgtgaaga tggtcagctg ctgggtagct gtattctgtt tcattttgaa
      661 aaaggtagcc gtcgtgcaga actgggttat tgtctggcac gtgcagcaca aggtcgtggt
      721 tatatgggtg aagcactgcg tcgtctgctg gcatttgcat ttgatgaaat tgatctgaat
      781 cgtctggaag cagaaatcga tccgcgtaat cgtccgagcg cagcaagcct ggaacgtctg
      841 ggttttcgtc aagaaggcct gctggcacag cgttggattg ttagcggtga agttagcgat
      901 agcgcactgt atggactgct ggcagaacat tggcgtaatc gctaactcga gcaccaccac
      961 caccaccact gagatccggc tgctaacaaa gcccgaaagg aagctgagtt ggctgctgcc
     1021 accgctgagc aataactagc ataacccctt ggggcctcta aacgggtctt gaggggtttt
     1081 ttgctgaaag gaggaactat atccggattg gcgaatggga cgcgccctgt agcggcgcat
     1141 taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag
     1201 cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc
     1261 aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc
     1321 ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga tagacggttt
     1381 ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa
     1441 caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg ccgatttcgg
     1501 cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat
     1561 taacgtttac aatttcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt
     1621 tatttttcta aatacattca aatatgtatc cgctcatgaa ttaattctta gaaaaactca
     1681 tcgagcatca aatgaaactg caatttattc atatcaggat tatcaatacc atatttttga
     1741 aaaagccgtt tctgtaatga aggagaaaac tcaccgaggc agttccatag gatggcaaga
     1801 tcctggtatc ggtctgcgat tccgactcgt ccaacatcaa tacaacctat taatttcccc
     1861 tcgtcaaaaa taaggttatc aagtgagaaa tcaccatgag tgacgactga atccggtgag
     1921 aatggcaaaa gtttatgcat ttctttccag acttgttcaa caggccagcc attacgctcg
     1981 tcatcaaaat cactcgcatc aaccaaaccg ttattcattc gtgattgcgc ctgagcgaga
     2041 cgaaatacgc gatcgctgtt aaaaggacaa ttacaaacag gaatcgaatg caaccggcgc
     2101 aggaacactg ccagcgcatc aacaatattt tcacctgaat caggatattc ttctaatacc
     2161 tggaatgctg ttttcccggg gatcgcagtg gtgagtaacc atgcatcatc aggagtacgg
     2221 ataaaatgct tgatggtcgg aagaggcata aattccgtca gccagtttag tctgaccatc
     2281 tcatctgtaa catcattggc aacgctacct ttgccatgtt tcagaaacaa ctctggcgca
     2341 tcgggcttcc catacaatcg atagattgtc gcacctgatt gcccgacatt atcgcgagcc
     2401 catttatacc catataaatc agcatccatg ttggaattta atcgcggcct agagcaagac
     2461 gtttcccgtt gaatatggct cataacaccc cttgtattac tgtttatgta agcagacagt
     2521 tttattgttc atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc
     2581 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt
     2641 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac
     2701 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg tccttctagt
     2761 gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
     2821 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
     2881 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
     2941 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
     3001 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
     3061 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
     3121 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
     3181 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
     3241 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc
     3301 ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag
     3361 cgaggaagcg gaagagcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc
     3421 acaccgcata tatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag
     3481 tatacactcc gctatcgcta cgtgactggg tcatggctgc gccccgacac ccgccaacac
     3541 ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga
     3601 ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgaggc
     3661 agctgcggta aagctcatca gcgtggtcgt gaagcgattc acagatgtct gcctgttcat
     3721 ccgcgtccag ctcgttgagt ttctccagaa gcgttaatgt ctggcttctg ataaagcggg
     3781 ccatgttaag ggcggttttt tcctgtttgg tcactgatgc ctccgtgtaa gggggatttc
     3841 tgttcatggg ggtaatgata ccgatgaaac gagagaggat gctcacgata cgggttactg
     3901 atgatgaaca tgcccggtta ctggaacgtt gtgagggtaa acaactggcg gtatggatgc
     3961 ggcgggacca gagaaaaatc actcagggtc aatgccagcg cttcgttaat acagatgtag
     4021 gtgttccaca gggtagccag cagcatcctg cgatgcagat ccggaacata atggtgcagg
     4081 gcgctgactt ccgcgtttcc agactttacg aaacacggaa accgaagacc attcatgttg
     4141 ttgctcaggt cgcagacgtt ttgcagcagc agtcgcttca cgttcgctcg cgtatcggtg
     4201 attcattctg ctaaccagta aggcaacccc gccagcctag ccgggtcctc aacgacagga
     4261 gcacgatcat gcgcacccgt ggggccgcca tgccggcgat aatggcctgc ttctcgccga
     4321 aacgtttggt ggcgggacca gtgacgaagg cttgagcgag ggcgtgcaag attccgaata
     4381 ccgcaagcga caggccgatc atcgtcgcgc tccagcgaaa gcggtcctcg ccgaaaatga
     4441 cccagagcgc tgccggcacc tgtcctacga gttgcatgat aaagaagaca gtcataagtg
     4501 cggcgacgat agtcatgccc cgcgcccacc ggaaggagct gactgggttg aaggctctca
     4561 agggcatcgg tcgagatccc ggtgcctaat gagtgagcta acttacatta attgcgttgc
     4621 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
     4681 aacgcgcggg gagaggcggt ttgcgtattg ggcgccaggg tggtttttct tttcaccagt
     4741 gagacgggca acagctgatt gcccttcacc gcctggccct gagagagttg cagcaagcgg
     4801 tccacgctgg tttgccccag caggcgaaaa tcctgtttga tggtggttaa cggcgggata
     4861 taacatgagc tgtcttcggt atcgtcgtat cccactaccg agatatccgc accaacgcgc
     4921 agcccggact cggtaatggc gcgcattgcg cccagcgcca tctgatcgtt ggcaaccagc
     4981 atcgcagtgg gaacgatgcc ctcattcagc atttgcatgg tttgttgaaa accggacatg
     5041 gcactccagt cgccttcccg ttccgctatc ggctgaattt gattgcgagt gagatattta
     5101 tgccagccag ccagacgcag acgcgccgag acagaactta atgggcccgc taacagcgcg
     5161 atttgctggt gacccaatgc gaccagatgc tccacgccca gtcgcgtacc gtcttcatgg
     5221 gagaaaataa tactgttgat gggtgtctgg tcagagacat caagaaataa cgccggaaca
     5281 ttagtgcagg cagcttccac agcaatggca tcctggtcat ccagcggata gttaatgatc
     5341 agcccactga cgcgttgcgc gagaagattg tgcaccgccg ctttacaggc ttcgacgccg
     5401 cttcgttcta ccatcgacac caccacgctg gcacccagtt gatcggcgcg agatttaatc
     5461 gccgcgacaa tttgcgacgg cgcgtgcagg gccagactgg aggtggcaac gccaatcagc
     5521 aacgactgtt tgcccgccag ttgttgtgcc acgcggttgg gaatgtaatt cagctccgcc
     5581 atcgccgctt ccactttttc ccgcgttttc gcagaaacgt ggctggcctg gttcaccacg
     5641 cgggaaacgg tctgataaga gacaccggca tactctgcga catcgtataa cgttactggt
     5701 ttcacattca ccaccctgaa ttgactctct tccgggcgct atcatgccat accgcgaaag
     5761 gttttgcgcc attcgatggt gtccgggatc tcgacgctct cccttatgcg actcctgcat
     5821 taggaagcag cccagtagta ggttgagg
//