LOCUS       Expected_topocloning_pr 4304 bp ds-DNA     circular     22-OCT-2022
DEFINITION  .
KEYWORDS    "creator:Thermo Fisher (Invitrogen)" "marker:NeoR/KanR, AmpR"
            "organism:Escherichia coli"
COMMENT     Linearized vector with bound topoisomerase, for TOPO® cloning of
            blunt PCR products. Can be used to generate nested deletions for
            sequencing. Do not use phosphorylated primers for the PCR
            amplification because 5' phosphates will prevent ligation into this
            vector.
FEATURES             Location/Qualifiers
     rep_origin      1..589
                     /label="ori"
                     /ApEinfo_revcolor="#ffef86"
                     /ApEinfo_fwdcolor="#ffef86"
                     /note="direction: RIGHT note: high-copy-number ColE1/pMB1/pBR322/pUC origin of replication"
     CDS             987..1064
                     /label="lacZα"
                     /ApEinfo_revcolor="#993366"
                     /ApEinfo_fwdcolor="#993366"
                     /note="codon_start: 1 gene: lacZ fragment product: LacZα fragment of β-galactosidase transl_table: 1 translation: MTMITPSSELTLTKGTSPAGLNEFAL"
     promoter        1012..1030
                     /label="T3 promoter"
                     /ApEinfo_revcolor="#c6c9d1"
                     /ApEinfo_fwdcolor="#c6c9d1"
                     /note="note: promoter for bacteriophage T3 RNA polymerase"
     misc_feature    1065..1093
                     /label="illumina P5"
                     /ApEinfo_revcolor="#b4abac"
                     /ApEinfo_fwdcolor="#b4abac"
     misc_feature    1094..1101
                     /label="i5"
                     /ApEinfo_revcolor="#f58a5e"
                     /ApEinfo_fwdcolor="#f58a5e"
     misc_feature    1102..1134
                     /label="Nextera R1"
                     /ApEinfo_revcolor="#ff9ccd"
                     /ApEinfo_fwdcolor="#ff9ccd"
     misc_feature    1135..1234
                     /label="any cDNA"
                     /ApEinfo_revcolor="#ffef86"
                     /ApEinfo_fwdcolor="#ffef86"
     misc_feature    1235..1245
                     /label="polyA"
                     /ApEinfo_revcolor="#d6b295"
                     /ApEinfo_fwdcolor="#d6b295"
     misc_feature    1246..1255
                     /label="UMI"
                     /ApEinfo_revcolor="#faac61"
                     /ApEinfo_fwdcolor="#faac61"
     misc_feature    1256..1289
                     /label="Nextera R2"
                     /ApEinfo_revcolor="#ff9ccd"
                     /ApEinfo_fwdcolor="#ff9ccd"
     misc_feature    1290..1388
                     /label="cell barcode"
                     /ApEinfo_revcolor="#faac61"
                     /ApEinfo_fwdcolor="#faac61"
     misc_feature    complement(1389..1412)
                     /label="illumina P7"
                     /ApEinfo_revcolor="#c6c9d1"
                     /ApEinfo_fwdcolor="#c6c9d1"
     CDS             1413..1928
                     /label="lacZα"
                     /ApEinfo_revcolor="#993366"
                     /ApEinfo_fwdcolor="#993366"
                     /note="codon_start: 1 gene: lacZ fragment product: LacZα fragment of β-galactosidase transl_table: 1 translation: KGEFAAAKFNSPYSESYYNSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEARTDRPSQQLRSLYVRQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDKVSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI*"
     promoter        complement(1447..1465)
                     /label="T7 promoter"
                     /ApEinfo_revcolor="#c6c9d1"
                     /ApEinfo_fwdcolor="#c6c9d1"
                     /note="note: promoter for bacteriophage T7 RNA polymerase"
     CDS             2277..3071
                     /label="NeoR/KanR"
                     /ApEinfo_revcolor="#b7e6d7"
                     /ApEinfo_fwdcolor="#b7e6d7"
                     /note="codon_start: 1 gene: aph(3')-II (or nptII) note: confers resistance to neomycin, kanamycin, and G418 (Geneticin®) product: aminoglycoside phosphotransferase from Tn5 transl_table: 1 translation: MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQGLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF*"
     CDS             complement(3321..4112)
                     /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(4113..4181)
                     /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*"
ORIGIN
        1 TTGAGATCCT TTTTTTCTGC GCGTAATCTG CTGCTTGCAA ACAAAAAAAC CACCGCTACC
       61 AGCGGTGGTT TGTTTGCCGG ATCAAGAGCT ACCAACTCTT TTTCCGAAGG TAACTGGCTT
      121 CAGCAGAGCG CAGATACCAA ATACTGTTCT TCTAGTGTAG CCGTAGTTAG GCCACCACTT
      181 CAAGAACTCT GTAGCACCGC CTACATACCT CGCTCTGCTA ATCCTGTTAC CAGTGGCTGC
      241 TGCCAGTGGC GATAAGTCGT GTCTTACCGG GTTGGACTCA AGACGATAGT TACCGGATAA
      301 GGCGCAGCGG TCGGGCTGAA CGGGGGGTTC GTGCACACAG CCCAGCTTGG AGCGAACGAC
      361 CTACACCGAA CTGAGATACC TACAGCGTGA GCTATGAGAA AGCGCCACGC TTCCCGAAGG
      421 GAGAAAGGCG GACAGGTATC CGGTAAGCGG CAGGGTCGGA ACAGGAGAGC GCACGAGGGA
      481 GCTTCCAGGG GGAAACGCCT GGTATCTTTA TAGTCCTGTC GGGTTTCGCC ACCTCTGACT
      541 TGAGCGTCGA TTTTTGTGAT GCTCGTCAGG GGGGCGGAGC CTATGGAAAA ACGCCAGCAA
      601 CGCGGCCTTT TTACGGTTCC TGGCCTTTTG CTGGCCTTTT GCTCACATGT TCTTTCCTGC
      661 GTTATCCCCT GATTCTGTGG ATAACCGTAT TACCGCCTTT GAGTGAGCTG ATACCGCTCG
      721 CCGCAGCCGA ACGACCGAGC GCAGCGAGTC AGTGAGCGAG GAAGCGGAAG AGCGCCCAAT
      781 ACGCAAACCG CCTCTCCCCG CGCGTTGGCC GATTCATTAA TGCAGCTGGC ACGACAGGTT
      841 TCCCGACTGG AAAGCGGGCA GTGAGCGCAA CGCAATTAAT GTGAGTTAGC TCACTCATTA
      901 GGCACCCCAG GCTTTACACT TTATGCTTCC GGCTCGTATG TTGTGTGGAA TTGTGAGCGG
      961 ATAACAATTT CACACAGGAA ACAGCTATGA CCATGATTAC GCCAAGCTCA GAATTAACCC
     1021 TCACTAAAGG GACTAGTCCT GCAGGTTTAA ACGAATTCGC CCTTAATGAT ACGGCGACCA
     1081 CCGAGATCTA CACNNNNNNN NTCGTCGGCA GCGTCAGATG TGTATAAGAG ACAGNNNNNN
     1141 NNNNNNNNNN NNNNNNNNNN NNNNNNNNNN NNNNNNNNNN NNNNNNNNNN NNNNNNNNNN
     1201 NNNNNNNNNN NNNNNNNNNN NNNNNNNNNN NNNNAAAAAA AAAAANNNNN NNNNNCTGTC
     1261 TCTTATACAC ATCTCCGAGC CCACGAGACT CGGACGATCA TGGGNNNNNN NNCAAGTATG
     1321 CAGCGCGCTC AAGCACGTGG ATNNNNNNNN AGTCGTACGC CGATGCGAAA CATCGGCCAC
     1381 NNNNNNNNAT CTCGTATGCC GTCTTCTGCT TGAAGGGCGA ATTCGCGGCC GCTAAATTCA
     1441 ATTCGCCCTA TAGTGAGTCG TATTACAATT CACTGGCCGT CGTTTTACAA CGTCGTGACT
     1501 GGGAAAACCC TGGCGTTACC CAACTTAATC GCCTTGCAGC ACATCCCCCT TTCGCCAGCT
     1561 GGCGTAATAG CGAAGAGGCC CGCACCGATC GCCCTTCCCA ACAGTTGCGC AGCCTATACG
     1621 TACGGCAGTT TAAGGTTTAC ACCTATAAAA GAGAGAGCCG TTATCGTCTG TTTGTGGATG
     1681 TACAGAGTGA TATTATTGAC ACGCCGGGGC GACGGATGGT GATCCCCCTG GCCAGTGCAC
     1741 GTCTGCTGTC AGATAAAGTC TCCCGTGAAC TTTACCCGGT GGTGCATATC GGGGATGAAA
     1801 GCTGGCGCAT GATGACCACC GATATGGCCA GTGTGCCGGT CTCCGTTATC GGGGAAGAAG
     1861 TGGCTGATCT CAGCCACCGC GAAAATGACA TCAAAAACGC CATTAACCTG ATGTTCTGGG
     1921 GAATATAAAT GTCAGGCATG AGATTATCAA AAAGGATCTT CACCTAGATC CTTTTCACGT
     1981 AGAAAGCCAG TCCGCAGAAA CGGTGCTGAC CCCGGATGAA TGTCAGCTAC TGGGCTATCT
     2041 GGACAAGGGA AAACGCAAGC GCAAAGAGAA AGCAGGTAGC TTGCAGTGGG CTTACATGGC
     2101 GATAGCTAGA CTGGGCGGTT TTATGGACAG CAAGCGAACC GGAATTGCCA GCTGGGGCGC
     2161 CCTCTGGTAA GGTTGGGAAG CCCTGCAAAG TAAACTGGAT GGCTTTCTTG CCGCCAAGGA
     2221 TCTGATGGCG CAGGGGATCA AGCTCTGATC AAGAGACAGG ATGAGGATCG TTTCGCATGA
     2281 TTGAACAAGA TGGATTGCAC GCAGGTTCTC CGGCCGCTTG GGTGGAGAGG CTATTCGGCT
     2341 ATGACTGGGC ACAACAGACA ATCGGCTGCT CTGATGCCGC CGTGTTCCGG CTGTCAGCGC
     2401 AGGGGCGCCC GGTTCTTTTT GTCAAGACCG ACCTGTCCGG TGCCCTGAAT GAACTGCAAG
     2461 ACGAGGCAGC GCGGCTATCG TGGCTGGCCA CGACGGGCGT TCCTTGCGCA GCTGTGCTCG
     2521 ACGTTGTCAC TGAAGCGGGA AGGGACTGGC TGCTATTGGG CGAAGTGCCG GGGCAGGATC
     2581 TCCTGTCATC TCACCTTGCT CCTGCCGAGA AAGTATCCAT CATGGCTGAT GCAATGCGGC
     2641 GGCTGCATAC GCTTGATCCG GCTACCTGCC CATTCGACCA CCAAGCGAAA CATCGCATCG
     2701 AGCGAGCACG TACTCGGATG GAAGCCGGTC TTGTCGATCA GGATGATCTG GACGAAGAGC
     2761 ATCAGGGGCT CGCGCCAGCC GAACTGTTCG CCAGGCTCAA GGCGAGCATG CCCGACGGCG
     2821 AGGATCTCGT CGTGACCCAT GGCGATGCCT GCTTGCCGAA TATCATGGTG GAAAATGGCC
     2881 GCTTTTCTGG ATTCATCGAC TGTGGCCGGC TGGGTGTGGC GGACCGCTAT CAGGACATAG
     2941 CGTTGGCTAC CCGTGATATT GCTGAAGAGC TTGGCGGCGA ATGGGCTGAC CGCTTCCTCG
     3001 TGCTTTACGG TATCGCCGCT CCCGATTCGC AGCGCATCGC CTTCTATCGC CTTCTTGACG
     3061 AGTTCTTCTG AATTATTAAC GCTTACAATT TCCTGATGCG GTATTTTCTC CTTACGCATC
     3121 TGTGCGGTAT TTCACACCGC ATCAGGTGGC ACTTTTCGGG GAAATGTGCG CGGAACCCCT
     3181 ATTTGTTTAT TTTTCTAAAT ACATTCAAAT ATGTATCCGC TCATGAGATT ATCAAAAAGG
     3241 ATCTTCACCT AGATCCTTTT AAATTAAAAA TGAAGTTTTA AATCAATCTA AAGTATATAT
     3301 GAGTAAACTT GGTCTGACAG TTACCAATGC TTAATCAGTG AGGCACCTAT CTCAGCGATC
     3361 TGTCTATTTC GTTCATCCAT AGTTGCCTGA CTCCCCGTCG TGTAGATAAC TACGATACGG
     3421 GAGGGCTTAC CATCTGGCCC CAGTGCTGCA ATGATACCGC GAGACCCACG CTCACCGGCT
     3481 CCAGATTTAT CAGCAATAAA CCAGCCAGCC GGAAGGGCCG AGCGCAGAAG TGGTCCTGCA
     3541 ACTTTATCCG CCTCCATCCA GTCTATTAAT TGTTGCCGGG AAGCTAGAGT AAGTAGTTCG
     3601 CCAGTTAATA GTTTGCGCAA CGTTGTTGCC ATTGCTACAG GCATCGTGGT GTCACGCTCG
     3661 TCGTTTGGTA TGGCTTCATT CAGCTCCGGT TCCCAACGAT CAAGGCGAGT TACATGATCC
     3721 CCCATGTTGT GCAAAAAAGC GGTTAGCTCC TTCGGTCCTC CGATCGTTGT CAGAAGTAAG
     3781 TTGGCCGCAG TGTTATCACT CATGGTTATG GCAGCACTGC ATAATTCTCT TACTGTCATG
     3841 CCATCCGTAA GATGCTTTTC TGTGACTGGT GAGTACTCAA CCAAGTCATT CTGAGAATAG
     3901 TGTATGCGGC GACCGAGTTG CTCTTGCCCG GCGTCAATAC GGGATAATAC CGCGCCACAT
     3961 AGCAGAACTT TAAAAGTGCT CATCATTGGA AAACGTTCTT CGGGGCGAAA ACTCTCAAGG
     4021 ATCTTACCGC TGTTGAGATC CAGTTCGATG TAACCCACTC GTGCACCCAA CTGATCTTCA
     4081 GCATCTTTTA CTTTCACCAG CGTTTCTGGG TGAGCAAAAA CAGGAAGGCA AAATGCCGCA
     4141 AAAAAGGGAA TAAGGGCGAC ACGGAAATGT TGAATACTCA TACTCTTCCT TTTTCAATAT
     4201 TATTGAAGCA TTTATCAGGG TTATTGTCTC ATGACCAAAA TCCCTTAACG TGAGTTTTCG
     4261 TTCCACTGAG CGTCAGACCC CGTAGAAAAG ATCAAAGGAT CTTC
//