LOCUS       sequence-316184-        4802 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  attB and attP-flanked pPgk-pEM7-Neo*-bpA positive selection 
            cassette.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4802)
  AUTHORS   Nam HS, Capecchi MR
  TITLE     Additional gene targeting vector construction plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 4802)
  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: 
            "Unpublished"
COMMENT     Sequence Label: attB Neo* attP
FEATURES             Location/Qualifiers
     source          1..4802
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..500
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(36..59)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     433..452
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     promoter        512..559
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             578..1381
                     /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="MGSAIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQ
                     GRPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQD
                     LLSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE
                     EHQGLAPAELFARLKARMPDGDDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQ
                     DIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(641..660)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1251..1270
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(1413..1430)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1419..1643
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    1701..1734
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="minimal attB site for the phi-C31 integrase (Groth 
                     et al., 2000)"
     primer_bind     complement(1755..1771)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1755..1771)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     promoter        complement(1808..1826)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1847..1863)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1847..1863)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1860..1882)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1871..1887
                     /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(1895..1901,1902..1919,1920..1925))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1940..1961
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2078..2095)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2249..2837)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2329..2348)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3008..3799,3800..3868))
                     /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     3631..3650
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3869..3973)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(3999..4454)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4136..4157)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4348..4367
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4581..4603
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4595..4612
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4596..4612
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4619..4638
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4619..4637
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     4662..4678
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     4663..4679
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 gggtagggga ggcgcttttc ccaaggcagt ctggagcatg cgctttagca gccccgctgg
       61 gcacttggcg ctacacaagt ggcctctggc ctcgcacaca ttccacatcc accggtaggc
      121 gccaaccggc tccgttcttt ggtggcccct tcgcgccacc ttctactcct cccctagtca
      181 ggaagttccc ccccgccccg cagctcgcgt cgtgcaggac gtgacaaatg gaagtagcac
      241 gtctcactag tctcgtgcag atggacagca ccgctgagca atggaagcgg gtaggccttt
      301 ggggcagcgg ccaatagcag ctttgctcct tcgctttctg ggctcagagg ctgggaaggg
      361 gtgggtccgg gggcgggctc aggggcgggc tcaggggcgg ggcgggcgcc cgaaggtcct
      421 ccggaggccc ggcattctgc acgcttcaaa agcgcacgtc tgccgcgctg ttctcctctt
      481 cctcatctcc gggcctttcg acctgcagcc tgttgacaat taatcatcgg catagtatat
      541 cggcatagta taatacgaca aggtgaggaa ctaaaccatg ggatcggcca ttgaacaaga
      601 tggattgcac gcaggttctc cggccgcttg ggtggagagg ctattcggct atgactgggc
      661 acaacagaca atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc
      721 ggttcttttt gtcaagaccg acctgtccgg tgccctgaat gaactgcagg acgaggcagc
      781 gcggctatcg tggctggcca cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac
      841 tgaagcggga agggactggc tgctattggg cgaagtgccg gggcaggatc tcctgtcatc
      901 tcaccttgct cctgccgaga aagtatccat catggctgat gcaatgcggc ggctgcatac
      961 gcttgatccg gctacctgcc cattcgacca ccaagcgaaa catcgcatcg agcgagcacg
     1021 tactcggatg gaagccggtc ttgtcgatca ggatgatctg gacgaagagc atcaggggct
     1081 cgcgccagcc gaactgttcg ccaggctcaa ggcgcgcatg cccgacggcg atgatctcgt
     1141 cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg
     1201 attcatcgac tgtggccggc tgggtgtggc ggaccgctat caggacatag cgttggctac
     1261 ccgtgatatt gctgaagagc ttggcggcga atgggctgac cgcttcctcg tgctttacgg
     1321 tatcgccgct cccgattcgc agcgcatcgc cttctatcgc cttcttgacg agttcttctg
     1381 aggggatcaa ttctctagag ctcgctgatc agcctcgact gtgccttcta gttgccagcc
     1441 atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt
     1501 cctttcctaa taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct
     1561 ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc
     1621 tggggatgcg gtgggctcta tggcttctga ggcggaaaga accagctggg gctcgactag
     1681 agcttgcgga acccttcgaa cgcgcccggg gagcccaagg gcacgccctg gcaccatcag
     1741 tcaggtacat aatggatcca ctagttctag agcggccgcc accgcggtgg agctccagct
     1801 tttgttccct ttagtgaggg ttaatttcga gcttggcgta atcatggtca tagctgtttc
     1861 ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt
     1921 gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc
     1981 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg
     2041 ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct
     2101 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca
     2161 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
     2221 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
     2281 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     2341 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     2401 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
     2461 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     2521 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     2581 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     2641 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
     2701 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     2761 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
     2821 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
     2881 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
     2941 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
     3001 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     3061 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     3121 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
     3181 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     3241 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     3301 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
     3361 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     3421 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     3481 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     3541 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     3601 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
     3661 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     3721 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     3781 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     3841 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     3901 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     3961 taggggttcc gcgcacattt ccccgaaaag tgccacctaa attgtaagcg ttaatatttt
     4021 gttaaaattc gcgttaaatt tttgttaaat cagctcattt tttaaccaat aggccgaaat
     4081 cggcaaaatc ccttataaat caaaagaata gaccgagata gggttgagtg ttgttccagt
     4141 ttggaacaag agtccactat taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt
     4201 ctatcagggc gatggcccac tacgtgaacc atcaccctaa tcaagttttt tggggtcgag
     4261 gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc cgatttagag cttgacgggg
     4321 aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg aaaggagcgg gcgctagggc
     4381 gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc
     4441 gctacagggc gcgtcccatt cgccattcag gctgcgcaac tgttgggaag ggcgatcggt
     4501 gcgggcctct tcgctattac gccagctggc gaaaggggga tgtgctgcaa ggcgattaag
     4561 ttgggtaacg ccagggtttt cccagtcacg acgttgtaaa acgacggcca gtgaattgta
     4621 atacgactca ctatagggcg aattgggtac cgggcccccc ctcgaggtcg acggtatcga
     4681 taagcttgat atcgaattcc cccccaactg agagaactca aaggttaccc cagttgggga
     4741 ggtctgaaga ggagtttacg tccagccaag ctagcttggc tgcaggtcgt cgaaattcta
     4801 cc
//
