LOCUS       sequence-429705-        5262 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Gateway Donor Vector with attP1 and attP2 sites modified from 
            PDONR221 and Ampicillin resistance.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5262)
  AUTHORS   Sivakumar P, Vaishnavi V, Gayatri K, Satheesh GR, Siddiqi I
  TITLE     Improved validation of protein interactions using bicistronic BiFC 
            (Bi2FC).
  JOURNAL   Physiol Mol Biol Plants. 2024 Jul;30(7):1047-1054. doi: 
            10.1007/s12298-024-01477-y. Epub 2024 Jul 2.
  PUBMED    39100877
REFERENCE   2  (bases 1 to 5262)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1007/s12298-024-01477-y"; journalName: "Physiol Mol Biol 
            Plants"; date: "2024-07"; volume: "30"; issue: "7"; pages: 
            "1047-1054"
COMMENT     Sequence Label: pCONR221
FEATURES             Location/Qualifiers
     source          1..5262
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     93..110
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        183..260
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     complement(280..299)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     terminator      complement(649..676)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(768..854)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     903..925
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     917..934
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     918..934
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    951..1182
                     /gene="mutant version of attP"
                     /label=attP1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             complement(1578..1883)
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     complement(1680..1699)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             complement(2203..2886)
                     /codon_start=1
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGRKRR
                     VDPAY"
     primer_bind     2801..2820
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(2887..2989)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     protein_bind    complement(3134..3365)
                     /gene="mutant version of attP"
                     /label=attP2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(3391..3409)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3477..3581
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3582..3650,3651..4442)
                     /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(3800..3819)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4613..5201
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5102..5121
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg
       61 ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag
      121 agcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcataaatt
      181 ccgacaccat cgaatggcgc aaaacctttc gcggtatggc atgatagcgc ccggaagaga
      241 gtcaattcag ggtggtgaat gtgaaaccag taacgttata cgatgtcgca gagtatgccg
      301 gtgtctctta tcagaccgtt tcccgcgtgg tgaaccaggc cagccacgtt tctgcgaaaa
      361 cgcgggaaaa agtggaagcg gcgatggcgg agctgaatta cattcccaac cgcgtggcac
      421 aacaactggc gggcaaacag tcgttgctga ttggcgttgc cacctccagt ctggccctgc
      481 acgcgccgtc gcaaattgtc gcggcgatta aatctcgcgc cgatcaactg ggtgccagcg
      541 tggtggtgtc gatggtagaa cgaagcggcg tcgaagcctg taaagcggcg gtgcacaatc
      601 ttctcgcgca acgcgtaccg ctagccagga agagtttgta gaaacgcaaa aaggccatcc
      661 gtcaggatgg ccttctgctt agtttgatgc ctggcagttt atggcgggcg tcctgcccgc
      721 caccctccgg gccgttgctt cacaacgttc aaatccgctc ccggcggatt tgtcctactc
      781 aggagagcgt tcaccgacaa acaacagata aaacgaaagg cccagtcttc cgactgagcc
      841 tttcgtttta tttgatgcct ggcagttccc tactctcgcg ttaacgctag catggatgtt
      901 ttcccagtca cgacgttgta aaacgacggc cagtcttaag ctcgggcccc aaataatgat
      961 tttattttga ctgatagtga cctgttcgtt gcaacaaatt gatgagcaat gcttttttat
     1021 aatgccaact ttgtacaaaa aagctgaacg agaaacgtaa aatgatataa atatcaatat
     1081 attaaattag attttgcata aaaaacagac tacataatac tgtaaaacac aacatatcca
     1141 gtcactatga atcaactact tagatggtat tagtgacctg tagtcgaccg acagccttcc
     1201 aaatgttctt cgggtgatgc tgccaactta gtcgaccgac agccttccaa atgttcttct
     1261 caaacggaat cgtcgtatcc agcctactcg ctattgtcct caatgccgta ttaaatcata
     1321 aaaagaaata agaaaaagag gtgcgagcct cttttttgtg tgacaaaata aaaacatcta
     1381 cctattcata tacgctagtg tcatagtcct gaaaatcatc tgcatcaaga acaatttcac
     1441 aactcttata cttttctctt acaagtcgtt cggcttcatc tggattttca gcctctatac
     1501 ttactaaacg tgataaagtt tctgtaattt ctactgtatc gacctgcaga ctggctgtgt
     1561 ataagggagc ctgacattta tattccccag aacatcaggt taatggcgtt tttgatgtca
     1621 ttttcgcggt ggctgagatc agccacttct tccccgataa cggagaccgg cacactggcc
     1681 atatcggtgg tcatcatgcg ccagctttca tccccgatat gcaccaccgg gtaaagttca
     1741 cgggagactt tatctgacag cagacgtgca ctggccaggg ggatcaccat ccgtcgcccg
     1801 ggcgtgtcaa taatatcact ctgtacatcc acaaacagac gataacggct ctctctttta
     1861 taggtgtaaa ccttaaactg catttcacca gcccctgttc tcgtcagcaa aagagccgtt
     1921 catttcaata aaccgggcga cctcagccat cccttcctga ttttccgctt tccagcgttc
     1981 ggcacgcaga cgacgggctt cattctgcat ggttgtgctt accagaccgg agatattgac
     2041 atcatatatg ccttgagcaa ctgatagctg tcgctgtcaa ctgtcactgt aatacgctgc
     2101 ttcatagcat acctcttttt gacatacttc gggtatacat atcagtatat attcttatac
     2161 cgcaaaaatc agcgcgcaaa tacgcatact gttatctggc ttttagtaag ccggatccac
     2221 gcggcgttta cgccccccct gccactcatc gcagtactgt tgtaattcat taagcattct
     2281 gccgacatgg aagccatcac aaacggcatg atgaacctga atcgccagcg gcatcagcac
     2341 cttgtcgcct tgcgtataat atttgcccat ggtgaaaacg ggggcgaaga agttgtccat
     2401 attggccacg tttaaatcaa aactggtgaa actcacccag ggattggctg agacgaaaaa
     2461 catattctca ataaaccctt tagggaaata ggccaggttt tcaccgtaac acgccacatc
     2521 ttgcgaatat atgtgtagaa actgccggaa atcgtcgtgg tattcactcc agagcgatga
     2581 aaacgtttca gtttgctcat ggaaaacggt gtaacaaggg tgaacactat cccatatcac
     2641 cagctcaccg tctttcattg ccatacggaa ttccggatga gcattcatca ggcgggcaag
     2701 aatgtgaata aaggccggat aaaacttgtg cttatttttc tttacggtct ttaaaaaggc
     2761 cgtaatatcc agctgaacgg tctggttata ggtacattga gcaactgact gaaatgcctc
     2821 aaaatgttct ttacgatgcc attgggatat atcaacggtg gtatatccag tgattttttt
     2881 ctccatttta gcttccttag ctcctgaaaa tctcgataac tcaaaaaata cgcccggtag
     2941 tgatcttatt tcattatggt gaaagttgga acctcttacg tgccgatcaa cgtctcattt
     3001 tcgccaaaag ttggcccagg gcttcccggt atcaacaggg acaccaggat ttatttattc
     3061 tgcgaagtga tcttccgtca caggtattta ttcggcgcaa agtgcgtcgg gtgatgctgc
     3121 caacttagtc gactacaggt cactaatacc atctaagtag ttgattcata gtgactggat
     3181 atgttgtgtt ttacagtatt atgtagtctg ttttttatgc aaaatctaat ttaatatatt
     3241 gatatttata tcattttacg tttctcgttc agctttcttg tacaaagttg gcattataag
     3301 aaagcattgc ttatcaattt gttgcaacga acaggtcact atcagtcaaa ataaaatcat
     3361 tatttgccat ccagctgata agacgaaagg gcctcgtgat acgcctattt ttataggtta
     3421 atgtcatgat aataatggtt tcttagacgt caggtggcac ttttcgggga aatgtgcgcg
     3481 gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat
     3541 aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt caacatttcc
     3601 gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa
     3661 cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac
     3721 tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga
     3781 tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtgttgac gccgggcaag
     3841 agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca
     3901 cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca
     3961 tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa
     4021 ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc
     4081 tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgcagca atggcaacaa
     4141 cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag
     4201 actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct
     4261 ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac
     4321 tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa
     4381 ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt
     4441 aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat
     4501 ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg
     4561 agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc
     4621 ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg
     4681 tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag
     4741 cgcagatacc aaatactgtc cttctagtgt agccgtagtt aggccaccac ttcaagaact
     4801 ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg
     4861 gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc
     4921 ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg
     4981 aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg
     5041 cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag
     5101 ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc
     5161 gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct
     5221 ttttacggtt cctggccttt tgctggcctt ttgctcacat gt
//
