LOCUS       sequence-269109-        4506 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Kanamycin selection marker flanked by ~200 bp of homology to 
            bacteriophage lambda. The selection marker will recombineer 
            21.3-22.8 kb away from the cosL DNA end of the phage genome..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4506)
  AUTHORS   Kim Y, de la Torre A, Leal AA, Finkelstein IJ
  TITLE     Efficient modification of lambda-DNA substrates for single-molecule 
            studies.
  JOURNAL   Sci Rep. 2017 May 18;7(1):2071. doi: 10.1038/s41598-017-01984-x.
  PUBMED    28522818
REFERENCE   2  (bases 1 to 4506)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Sci Rep."; 
            date: "2017-05-18"; volume: "7(1):2071. doi"; pages: " 
            10.1038/s41598-017-01984-x"
COMMENT     Sequence Label: pIF251
FEATURES             Location/Qualifiers
     source          1..4506
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     56..78
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     70..87
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     71..87
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     97..116
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        97..115
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    434..453
                     /label=lac operator (symmetric)
                     /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-beta-D-thiogalactopyranoside (IPTG). The 
                     symmetric lac operator was optimized for tight binding of 
                     lac repressor."
     CDS             705..1499
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(759..778)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1369..1388
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(1789..1809)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(1789..1807)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1828..1844)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1828..1844)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1841..1863)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1852..1868
                     /label=lac operator
                     /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-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(1876..1882,1883..1900,1901..1906))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1921..1942
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2059..2076)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2230..2818)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2310..2329)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2989..3780,3781..3849))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     3612..3631
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3850..3954)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(3980..4435)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4117..4138)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4329..4348
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg taacgccagg gttttcccag
       61 tcacgacgtt gtaaaacgac ggccagtgag cgcgcgtaat acgactcact atagggcgaa
      121 ttgggtacgg ccgtcaaggc caagcttccc gggatcttac gggaatgatc cagattttgc
      181 taccaccatg actaacgcgc ttgcgggtaa acaaccgaag aatgcgacac tgacggcgct
      241 ggcagggctt tccacggcga aaaataaatt accgtatttt gcggaaaatg atgccgccag
      301 cctgactgaa ctgactcagg ttggcaggga tattctggca aaaaattccg ttgcagatgt
      361 tcttgaatac cttggggccg gtgagaattc ggcctttccg gcaggtgcgg agcattgcta
      421 cggcgattcc tagaattgtg agcgctcaca attctagttg gagagccact gttcatttaa
      481 ataaagctct tcatgcatgc ggccgctctt cccatggtgc gatcgctctt cgggatttaa
      541 ataggtacct atggacagca agcgaaccgg aattgccagc tggggcgccc tctggtaagg
      601 ttgggaagcc ctgcaaagta aactggatgg ctttcttgcc gccaaggatc tgatggcgca
      661 ggggatcaag atctgatcaa gagacaggat gaggatcgtt tcgcatgatt gaacaagatg
      721 gattgcacgc aggttctccg gccgcttggg tggagaggct attcggctat gactgggcac
      781 aacagacaat cggctgctct gatgccgccg tgttccggct gtcagcgcag gggcgcccgg
      841 ttctttttgt caagaccgac ctgtccggtg ccctgaatga actgcaggac gaggcagcgc
      901 ggctatcgtg gctggccacg acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg
      961 aagcgggaag ggactggctg ctattgggcg aagtgccggg gcaggatctc ctgtcatctc
     1021 accttgctcc tgccgagaaa gtatccatca tggctgatgc aatgcggcgg ctgcatacgc
     1081 ttgatccggc tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta
     1141 ctcggatgga agccggtctt gtcgatcagg atgatctgga cgaagagcat caggggctcg
     1201 cgccagccga actgttcgcc aggctcaagg cgcgcatgcc cgacggcgag gatctcgtcg
     1261 tgacccatgg cgatgcctgc ttgccgaata tcatggtgga aaatggccgc ttttctggat
     1321 tcatcgactg tggccggctg ggtgtggcgg accgctatca ggacatagcg ttggctaccc
     1381 gtgatattgc tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg ctttacggta
     1441 tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct tcttgacgag ttcttctgat
     1501 ttcgtcatgt tttgagtctg ctgttgatat ttctaaagtc ggtttttttt cttcgttttc
     1561 tctaactatt ttccatgaaa tacatttttg attattattt gaatcaattc caattacctg
     1621 aagtctttca tctataattg gcattgtatg tattggttta ttggagtaga tgcttgcttt
     1681 tctgagccat agctctgata tccaaatgaa gccataggca tttgttattt tggctctgtc
     1741 agctgcatat aagtatcacg tgaagcttgc aagctccagc ttttgttccc tttagtgagg
     1801 gttaattgcg cgcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc
     1861 gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta
     1921 atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa
     1981 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat
     2041 tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg
     2101 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc
     2161 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
     2221 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag
     2281 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc
     2341 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
     2401 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
     2461 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
     2521 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
     2581 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
     2641 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa
     2701 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
     2761 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
     2821 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
     2881 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
     2941 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
     3001 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
     3061 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
     3121 gataccgcgt gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg
     3181 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
     3241 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
     3301 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
     3361 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt
     3421 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
     3481 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
     3541 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
     3601 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa
     3661 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta
     3721 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg
     3781 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg
     3841 aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat
     3901 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt
     3961 tccccgaaaa gtgccaccta aattgtaagc gttaatattt tgttaaaatt cgcgttaaat
     4021 ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat cccttataaa
     4081 tcaaaagaat agaccgagat agggttgagt gttgttccag tttggaacaa gagtccacta
     4141 ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca
     4201 ctacgtgaac catcacccta atcaagtttt ttggggtcga ggtgccgtaa agcactaaat
     4261 cggaacccta aagggagccc ccgatttaga gcttgacggg gaaagccggc gaacgtggcg
     4321 agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc
     4381 acgctgcgcg taaccaccac acccgccgcg cttaatgcgc cgctacaggg cgcgtcccat
     4441 tcgccattca ggctgcgcaa ctgttgggaa gggcgatcgg tgcgggcctc ttcgctatta
     4501 cgccag
//
