LOCUS       pJET1.2-tHe-Tub2D2      4485 bp ds-DNA     circular     26-APR-2023
DEFINITION  .
KEYWORDS    "accession:EF694056" "creator:Thermo Fisher (Fermentas)"
            "marker:AmpR" "organism:Escherichia coli"
COMMENT     Positive selection cloning vector with a lethal insert that allows
            for efficient recovery of blunt-ended PCR products. Linearize at
            Eco32I (EcoRV) to insert a blunt PCR product. colony 8D
FEATURES             Location/Qualifiers
     promoter          305..323
                       /label="T7 promoter"
                       /ApEinfo_revcolor="#c6c9d1"
                       /ApEinfo_fwdcolor="#c6c9d1"
                       /note="note: promoter for bacteriophage T7 RNA polymerase"
     misc_feature      352..1405
                       /label="8d-F-seq"
                       /ApEinfo_revcolor="#c6c9d1"
                       /ApEinfo_fwdcolor="#c6c9d1"
                       /note="verified by sequencing"
     primer            372..394
                       /label="HeTub2D2-FWD"
                       /note="sequence: ATGCTCGGACTCTCTACGCCTTC"
                       /ApEinfo_revcolor="#c6c9d1"
                       /ApEinfo_fwdcolor="#c6c9d1"
     CDS               372..1784
                       /label="tHe-Tub2D2"
                       /ApEinfo_revcolor="#ffef86"
                       /ApEinfo_fwdcolor="#ffef86"
     CDS               372..1784
                       /label="Translation 372-1784"
                       /translation="MLGLSTPSVLRLPVPSTMREIVHLQAGQCGNQIGSKFWETINGEHGIDPTGSYTGDNANQLERIDVYYQNASSGKYVPRAVLVDLEPGTMDSIRSSEYGQLFRPDNFVFGQSGAGNNWAKGHYTEGADLIDTVIDVVRKEAEHCDCLQGFQLTHSLGGGTGSGMGTLLISKIREEYPDRIMTTFSVVPSPKVSDTVVEPYNATLSVHQLVENTDETFCIDNEALYDICFRTLKLSTPAYGDLNHLVSATMSGVTTCFRFPGQLNADLRKLAVNMVPFPRLHFFMPGFAPLTARGSQTYRAMSVSELTQQMFDSKNMMTACDPRHGRYLTVAAIFRGRMSMKEVDDNMFNVQNKNSSHFVEWIPNNVKTAVCDIPPRGLSMSGTFIGNSTAIQEIFKRIGEQFTAMFRRKAFLHWYTGEGMDEMEFTEAESNMMDLVSEYQQYQEATVDDGAEFDEDTENVVVNGHENETA*"
     Possible mismatch 418..418
                       /label="Ambiguous site"
                       /ApEinfo_revcolor="#f8d3a9"
                       /ApEinfo_fwdcolor="#f8d3a9"
                       /note="sequencing chromatogram ambiguous might be also T - this would lead to Ser->Leu mutation"
     misc_feature      complement(891..1887)
                       /label="8d-R-seq"
                       /ApEinfo_revcolor="#75c6a9"
                       /ApEinfo_fwdcolor="#75c6a9"
                       /note="verified by sequencing"
     primer            complement(1862..1882)
                       /label="HeTub2D2-REV"
                       /note="sequence: tttcgaccagcggtgaagtta"
                       /ApEinfo_revcolor="#c6c9d1"
                       /ApEinfo_fwdcolor="#c6c9d1"
     protein_bind      2293..2309
                       /label="lac operator"
                       /ApEinfo_revcolor="#31849b"
                       /ApEinfo_fwdcolor="#31849b"
                       /note="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-β-D-thiogalactopyranoside (IPTG)."
     promoter          complement(2317..2323)
                       /label="lac UV5 promoter"
                       /ApEinfo_revcolor="#c6c9d1"
                       /ApEinfo_fwdcolor="#c6c9d1"
                       /note="note: E. coli lac promoter with an ""up"" mutation"
     promoter          complement(2324..2341)
                       /label="lac UV5 promoter"
                       /ApEinfo_revcolor="#c6c9d1"
                       /ApEinfo_fwdcolor="#c6c9d1"
                       /note="note: E. coli lac promoter with an ""up"" mutation"
     promoter          complement(2342..2347)
                       /label="lac UV5 promoter"
                       /ApEinfo_revcolor="#c6c9d1"
                       /ApEinfo_fwdcolor="#c6c9d1"
                       /note="note: E. coli lac promoter with an ""up"" mutation"
     rep_origin        complement(2674..3262)
                       /label="ori"
                       /ApEinfo_revcolor="#ffef86"
                       /ApEinfo_fwdcolor="#ffef86"
                       /note="direction: LEFT note: high-copy-number ColE1/pMB1/pBR322/pUC origin of replication"
     CDS               complement(3433..4224)
                       /label="AmpR"
                       /ApEinfo_revcolor="#b7e6d7"
                       /ApEinfo_fwdcolor="#b7e6d7"
                       /note="codon_start: 1 gene: bla note: confers resistance to ampicillin, carbenicillin, and related antibiotics product: β-lactamase transl_table: 1 translation: MSIQHFRVALIPFFAAFCLPVFA,HPETLVKVKDAEDQLGARVGYIELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRWEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGASLIKHW*"
     CDS               complement(4225..4293)
                       /label="AmpR"
                       /ApEinfo_revcolor="#b7e6d7"
                       /ApEinfo_fwdcolor="#b7e6d7"
                       /note="codon_start: 1 gene: bla note: confers resistance to ampicillin, carbenicillin, and related antibiotics product: β-lactamase transl_table: 1 translation: MSIQHFRVALIPFFAAFCLPVFA,HPETLVKVKDAEDQLGARVGYIELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRWEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGASLIKHW*"
     promoter          complement(4294..4398)
                       /label="AmpR promoter"
                       /ApEinfo_revcolor="#c6c9d1"
                       /ApEinfo_fwdcolor="#c6c9d1"
                       /note="gene: bla"
ORIGIN
        1 GCCCCTGCAG CCGAATTATA TTATTTTTGC CAAATAATTT TTAACAAAAG CTCTGAAGTC
       61 TTCTTCATTT AAATTCTTAG ATGATACTTC ATCTGGAAAA TTGTCCCAAT TAGTAGCATC
      121 ACGCTGTGAG TAAGTTCTAA ACCATTTTTT TATTGTTGTA TTATCTCTAA TCTTACTACT
      181 CGATGAGTTT TCGGTATTAT CTCTATTTTT AACTTGGAGC AGGTTCCATT CATTGTTTTT
      241 TTCATCATAG TGAATAAAAT CAACTGCTTT AACACTTGTG CCTGAACACC ATATCCATCC
      301 GGCGTAATAC GACTCACTAT AGGGAGAGCG GCCGCCAGAT CTTCCGGATG GCTCGAGTTT
      361 TTCAGCAAGA TATGCTCGGA CTCTCTACGC CTTCAGTACT GCGTCTTCCC GTTCCTTcGA
      421 CCATGCGAGA AATCGTCCAC CTCCAGGCCG GCCAATGCGG CAACCAAATC GGCTCCAAGT
      481 TCTGGGAAAC CATCAACGGC GAGCATGGAA TCGACCCCAC GGGATCTTAC ACGGGCGACA
      541 ACGCCAACCA GCTTGAGAGA ATCGATGTTT ACTACCAGAA TGCCAGCAGT GGGAAATATG
      601 TGCCAAGGGC CGTGTTGGTG GACCTTGAAC CGGGCACGAT GGACTCGATC CGGTCGTCCG
      661 AGTACGGCCA GCTCTTCCGT CCGGATAACT TTGTTTTCGG CCAAAGTGGG GCGGGCAACA
      721 ACTGGGCTAA GGGCCATTAC ACGGAGGGAG CCGACCTCAT CGACACGGTG ATCGATGTCG
      781 TCCGCAAGGA GGCCGAGCAC TGCGACTGCC TGCAGGGCTT CCAGCTGACC CACTCGCTCG
      841 GCGGTGGAAC CGGATCCGGC ATGGGCACCC TCCTCATCTC CAAGATCCGC GAGGAGTATC
      901 CCGATCGCAT CATGACCACA TTCAGTGTCG TTCCCTCGCC CAAGGTGTCC GACACCGTGG
      961 TGGAGCCGTA CAACGCCACC CTGTCAGTCC ACCAGCTGGT TGAGAACACC GACGAGACAT
     1021 TCTGCATCGA CAACGAGGCC CTCTACGACA TCTGCTTCCG CACCCTCAAG CTGTCAACCC
     1081 CGGCGTACGG CGACCTGAAC CATCTCGTCT CGGCGACCAT GTCCGGCGTG ACGACCTGCT
     1141 TcCGCTTcCC CGGCCAGTTG AACGCCGATC TCCGCAAACT GGCCGTCAAC ATGGTCCCCT
     1201 TCCCGCGGCT GCACTTCTTC ATGCCaGGCT TTGCGCCGCT GACTGCTCGC GGCTCTCAAA
     1261 CCTACCGCGC CATGTCCGTG TCGGAGTTGA CCCAGCAGAT GTTCGACTCG AAGAACATGA
     1321 TGACCGCCTG CGATCCCCGC CACGGACGCT ACCTCACCGT GGCTGCCATC TTCCGCGGCC
     1381 GGATGAGCAT GAAGGAGGTC GATGATAATA TGTTCAACGT GCAGAACAAA AATAGCAGCC
     1441 ATTTCGTTGA GTGGATCCCC AACAACGTCA AGACCGCCGT CTGCGATATC CCCCCGCGCG
     1501 GACTGTCCAT GTCGGGCACC TTCATCGGCA ACAGCACGGC CATTCAGGAG ATCTTCAAGC
     1561 GTATCGGCGA GCAGTTCACC GCCATGTTCC GCCGCAAGGC CTTCCTCCAT TGGTACACTG
     1621 GCGAAGGCAT GGACGAGATG GAATTCACCG AAGCCGAGTC CAACATGATG GACCTGGTGT
     1681 CGGAATACCA GCAGTACCAG GAGGCGACtG TTGATGACGG CGCGGAATTC GACGAGGATA
     1741 CGGAAAATGT CGTGGTCAAT GGACATGAAA ACGAGACCGC CTAGatggat Tattcctgca
     1801 ccaccgacct ctcacatcga gattgtggca ttgtttcagc tccctttttt gctgcagttt
     1861 ttaacttcac cgctggtcga aaATCTTTCT AGAAGATCTC CTACAATATT CTCAGCTGCC
     1921 ATGGAAAATC GATGTTCTTC TTTTATTCTC TCAAGATTTT CAGGCTGTAT ATTAAAACTT
     1981 ATATTAAGAA CTATGCTAAC CACCTCATCA GGAACCGTTG TAGGTGGCGT GGGTTTTCTT
     2041 GGCAATCGAC TCTCATGAAA ACTACGAGCT AAATATTCAA TATGTTCCTC TTGACCAACT
     2101 TTATTCTGCA TTTTTTTTGA ACGAGGTTTA GAGCAAGCTT CAGGAAACTG AGACAGGAAT
     2161 TTTATTAAAA ATTTAAATTT TGAAGAAAGT TCAGGGTTAA TAGCATCCAT TTTTTGCTTT
     2221 GCAAGTTCCT CAGCATTCTT AACAAAAGAC GTCTCTTTTG ACATGTTTAA AGTTTAAACC
     2281 TCCTGTGTGA AATTGTTATC CGCTCACAAT TCCACACATT ATACGAGCCG GAAGCATAAA
     2341 GTGTAAAGCC TGGGGTGCCT AATGAGTGAG CTAACTCACA TTAATTGCGT TGCGCTCACT
     2401 GCCAATTGCT TTCCAGTCGG GAAACCTGTC GTGCCAGCTG CATTAATGAA TCGGCCAACG
     2461 CGCGGGGAGA GGCGGTTTGC GTATTGGGCG CTCTTCCGCT TCCTCGCTCA CTGACTCGCT
     2521 GCGCTCGGTC GTTCGGCTGC GGCGAGCGGT ATCAGCTCAC TCAAAGGCGG TAATACGGTT
     2581 ATCCACAGAA TCAGGGGATA ACGCAGGAAA GAACATGTGA GCAAAAGGCC AGCAAAAGGC
     2641 CAGGAACCGT AAAAAGGCCG CGTTGCTGGC GTTTTTCCAT AGGCTCCGCC CCCCTGACGA
     2701 GCATCACAAA AATCGACGCT CAAGTCAGAG GTGGCGAAAC CCGACAGGAC TATAAAGATA
     2761 CCAGGCGTTT CCCCCTGGAA GCTCCCTCGT GCGCTCTCCT GTTCCGACCC TGCCGCTTAC
     2821 CGGATACCTG TCCGCCTTTC TCCCTTCGGG AAGCGTGGCG CTTTCTCATA GCTCACGCTG
     2881 TAGGTATCTC AGTTCGGTGT AGGTCGTTCG CTCCAAGCTG GGCTGTGTGC ACGAACCCCC
     2941 CGTTCAGCCC GACCGCTGCG CCTTATCCGG TAACTATCGT CTTGAGTCCA ACCCGGTAAG
     3001 ACACGACTTA TCGCCACTGG CAGCAGCCAC TGGTAACAGG ATTAGCAGAG CGAGGTATGT
     3061 AGGCGGTGCT ACAGAGTTCT TGAAGTGGTG GCCTAACTAC GGCTACACTA GAAGGACAGT
     3121 ATTTGGTATC TGCGCTCTGC TGAAGCCAGT TACCTTCGGA AAAAGAGTTG GTAGCTCTTG
     3181 ATCCGGCAAA CAAACCACCG CTGGTAGCGG TGGTTTTTTT GTTTGCAAGC AGCAGATTAC
     3241 GCGCAGAAAA AAAGGATCTC AAGAAGATCC TTTGATCTTT TCTACGGGGT CTGACGCTCA
     3301 GTGGAACGAA AACTCACGTT AAGGGATTTT GGTCATGAGA TTATCAAAAA GGATCTTCAC
     3361 CTAGATCCTT TTAAATTAAA AATGAAGTTT TAAATCAATC TAAAGTATAT ATGAGTAAAC
     3421 TTGGTCTGAC AGTTACCAAT GCTTAATCAG TGAGGCACCT ATCTCAGCGA TCTGTCTATT
     3481 TCGTTCATCC ATAGTTGCCT GACTCCCCGT CGTGTAGATA ACTACGATAC GGGAGGGCTT
     3541 ACCATCTGGC CCCAGTGCTG CAATGATACC GCGAGACCCA CGCTCACCGG CTCCAGATTT
     3601 ATCAGCAATA AACCAGCCAG CCGGAAGGGC CGAGCGCAGA AGTGGTCCTG CAACTTTATC
     3661 CGCCTCCATC CAGTCTATTA ATTGTTGCCG GGAAGCTAGA GTAAGTAGTT CGCCAGTTAA
     3721 TAGTTTGCGC AACGTTGTTG CCATTGCTAC AGGCATCGTG GTGTCACGCT CGTCGTTTGG
     3781 TATGGCTTCA TTCAGCTCCG GTTCCCAACG ATCAAGGCGA GTTACATGAT CCCCCATGTT
     3841 GTGCAAAAAA GCGGTTAGCT CCTTCGGTCC TCCGATCGTT GTCAGAAGTA AGTTGGCCGC
     3901 AGTGTTATCA CTCATGGTTA TGGCAGCACT GCATAATTCT CTTACTGTCA TGCCATCCGT
     3961 AAGATGCTTT TCTGTGACTG GTGAGTACTC AACCAAGTCA TTCTGAGAAT AGTGTATGCG
     4021 GCGACCGAGT TGCTCTTGCC CGGCGTCAAT ACGGGATAAT ACCGCGCCAC ATAGCAGAAC
     4081 TTTAAAAGTG CTCATCATTG GAAAACGTTC TTCGGGGCGA AAACTCTCAA GGATCTTACC
     4141 GCTGTTGAGA TCCAGTTCGA TGTAACCCAC TCGTGCACCC AACTGATCTT CAGCATCTTT
     4201 TACTTTCACC AGCGTTTCTG GGTGAGCAAA AACAGGAAGG CAAAATGCCG CAAAAAAGGG
     4261 AATAAGGGCG ACACGGAAAT GTTGAATACT CATACTCTTC CTTTTTCAAT ATTATTGAAG
     4321 CATTTATCAG GGTTATTGTC TCATGAGCGG ATACATATTT GAATGTATTT AGAAAAATAA
     4381 ACAAATAGGG GTTCCGCGCA CATTTCCCCG AAAAGTGCCA CCTGACGTCT AAGAAACCAT
     4441 TATTATCATG ACATTAACCT ATAAAAATAG GCGTATCACG AGGCC
//