LOCUS       sequence-242276-        6685 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for in vivo enhancer reporter assay. Plasmid contains a 
            beta-globin minimal promoter followed by a lacZ reporter gene 
            derived from pRS16, with the entire reporter cassette flanked by 
            attB sites..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6685)
  AUTHORS   Castro JP, Yancoskie MN, Marchini M, Belohlavy S, Hiramatsu L, Kucka
            M, Beluch WH, Naumann R, Skuplik I, Cobb J, Barton NH, Rolian C, 
            Chan YF
  TITLE     An integrative genomic analysis of the Longshanks selection 
            experiment for longer limbs in mice.
  JOURNAL   Elife. 2019 Jun 6;8:e42014. doi: 10.7554/eLife.42014.
  PUBMED    31169497
REFERENCE   2  (bases 1 to 6685)
  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: "Elife."; 
            date: "2019-06-6"; volume: "8:e42014. doi"; pages: " 
            10.7554/eLife.42014"
COMMENT     Sequence Label: pBeta-lacZ-attBx2
FEATURES             Location/Qualifiers
     source          1..6685
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     24..40
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     25..41
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             183..3230
                     /codon_start=1
                     /gene="lacZ"
                     /product="beta-galactosidase"
                     /label=lacZ
                     /translation="VVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEARTDRPSQQLRS
                     LNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNVTYPITVNPP
                     FVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVNSAFHLWCNGRWVGYGQDSRLPSEF
                     DLSAFLRAGENRLAVMVLRWSDGSYLEDQDMWRMSGIFRDVSLLHKPTTQISDFHVATR
                     FNDDFSRAVLEAEVQMCGELRDYLRVTVSLWQGETQVASGTAPFGGEIIDERGGYADRV
                     TLRLNVENPKLWSAEIPNLYRAVVELHTADGTLIEAEACDVGFREVRIENGLLLLNGKP
                     LLIRGVNRHEHHPLHGQVMDEQTMVQDILLMKQNNFNAVRCSHYPNHPLWYTLCDRYGL
                     YVVDEANIETHGMVPMNRLTDDPRWLPAMSERVTRMVQRDRNHPSVIIWSLGNESGHGA
                     NHDALYRWIKSVDPSRPVQYEGGGADTTATDIICPMYARVDEDQPFPAVPKWSIKKWLS
                     LPGETRPLILCEYAHAMGNSLGGFAKYWQAFRQYPRLQGGFVWDWVDQSLIKYDENGNP
                     WSAYGGDFGDTPNDRQFCMNGLVFADRTPHPALTEAKHQQQFFQFRLSGQTIEVTSEYL
                     FRHSDNELLHWMVALDGKPLASGEVPLDVAPQGKQLIELPELPQPESAGQLWLTVRVVQ
                     PNATAWSEAGHISAWQQWRLAENLSVTLPAASHAIPHLTTSEMDFCIELGNKRWQFNRQ
                     SGFLSQMWIGDKKQLLTPLRDQFTRAPLDNDIGVSEATRIDPNAWVERWKAAGHYQAEA
                     ALLQCTADTLADAVLITTAHAWQHQGKTLFISRKTYRIDGSGQMAITVDVEVASDTPHP
                     ARIGLNCQLAQVAERVNWLGLGPQENYPDRLTAACFDRWDLPLSDMYTPYVFPSENGLR
                     CGTRELNYGPHQWRGDFQFNISRYSQQQLMETSHRHLLHAEEGTWLNIDGFHMGIGGDD
                     SWSPSVSAEFQLSAGRYHYQLVWCQK"
     polyA_signal    3494..3628
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3531..3550)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3585..3604
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3636..3652)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3636..3652)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     protein_bind    3653..3722
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="attB site for the phi-C31 integrase (Groth et al., 
                     2000)"
     primer_bind     complement(3759..3779)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3759..3777)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3798..3814)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3798..3814)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3811..3833)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3822..3838
                     /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(3846..3852,3853..3870,3871..3876))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3891..3912
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4029..4046)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4200..4788)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4280..4299)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4959..5750,5751..5819))
                     /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     5582..5601
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5820..5924)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5950..6405)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6087..6108)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6299..6318
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6546..6563
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6547..6563
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6573..6592
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6573..6591
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    6616..6685
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="attB site for the phi-C31 integrase (Groth et al., 
                     2000)"
ORIGIN
        1 ggtaccgatt accgggcccc ccctcgaggt cgacggtatc gataagcttg atatcgaatt
       61 cctgcagccc gggctgggca taaaagtcag ggcagagcca tctattgctt acatttgctt
      121 ctagcctgca ggtcgaggag cgcagccttc cagaagcaga gcgcggcgcc atgggggatc
      181 ccgtcgtttt acaacgtcgt gactgggaaa accctggcgt tacccaactt aatcgccttg
      241 cagcacatcc ccctttcgcc agctggcgta atagcgaaga ggcccgcacc gatcgccctt
      301 cccaacagtt gcgcagcctg aatggcgaat ggcgctttgc ctggtttccg gcaccagaag
      361 cggtgccgga aagctggctg gagtgcgatc ttcctgaggc cgatactgtc gtcgtcccct
      421 caaactggca gatgcacggt tacgatgcgc ccatctacac caacgtgacc tatcccatta
      481 cggtcaatcc gccgtttgtt cccacggaga atccgacggg ttgttactcg ctcacattta
      541 atgttgatga aagctggcta caggaaggcc agacgcgaat tatttttgat ggcgttaact
      601 cggcgtttca tctgtggtgc aacgggcgct gggtcggtta cggccaggac agtcgtttgc
      661 cgtctgaatt tgacctgagc gcatttttac gcgccggaga aaaccgcctc gcggtgatgg
      721 tgctgcgctg gagtgacggc agttatctgg aagatcagga tatgtggcgg atgagcggca
      781 ttttccgtga cgtctcgttg ctgcataaac cgactacaca aatcagcgat ttccatgttg
      841 ccactcgctt taatgatgat ttcagccgcg ctgtactgga ggctgaagtt cagatgtgcg
      901 gcgagttgcg tgactaccta cgggtaacag tttctttatg gcagggtgaa acgcaggtcg
      961 ccagcggcac cgcgcctttc ggcggtgaaa ttatcgatga gcgtggtggt tatgccgatc
     1021 gcgtcacact acgtctgaac gtcgaaaacc cgaaactgtg gagcgccgaa atcccgaatc
     1081 tctatcgtgc ggtggttgaa ctgcacaccg ccgacggcac gctgattgaa gcagaagcct
     1141 gcgatgtcgg tttccgcgag gtgcggattg aaaatggtct gctgctgctg aacggcaagc
     1201 cgttgctgat tcgaggcgtt aaccgtcacg agcatcatcc tctgcatggt caggtcatgg
     1261 atgagcagac gatggtgcag gatatcctgc tgatgaagca gaacaacttt aacgccgtgc
     1321 gctgttcgca ttatccgaac catccgctgt ggtacacgct gtgcgaccgc tacggcctgt
     1381 atgtggtgga tgaagccaat attgaaaccc acggcatggt gccaatgaat cgtctgaccg
     1441 atgatccgcg ctggctaccg gcgatgagcg aacgcgtaac gcgaatggtg cagcgcgatc
     1501 gtaatcaccc gagtgtgatc atctggtcgc tggggaatga atcaggccac ggcgctaatc
     1561 acgacgcgct gtatcgctgg atcaaatctg tcgatccttc ccgcccggtg cagtatgaag
     1621 gcggcggagc cgacaccacg gccaccgata ttatttgccc gatgtacgcg cgcgtggatg
     1681 aagaccagcc cttcccggct gtgccgaaat ggtccatcaa aaaatggctt tcgctacctg
     1741 gagagacgcg cccgctgatc ctttgcgaat acgcccacgc gatgggtaac agtcttggcg
     1801 gtttcgctaa atactggcag gcgtttcgtc agtatccccg tttacagggc ggcttcgtct
     1861 gggactgggt ggatcagtcg ctgattaaat atgatgaaaa cggcaacccg tggtcggctt
     1921 acggcggtga ttttggcgat acgccgaacg atcgccagtt ctgtatgaac ggtctggtct
     1981 ttgccgaccg cacgccgcat ccagcgctga cggaagcaaa acaccagcag cagtttttcc
     2041 agttccgttt atccgggcaa accatcgaag tgaccagcga atacctgttc cgtcatagcg
     2101 ataacgagct cctgcactgg atggtggcgc tggatggtaa gccgctggca agcggtgaag
     2161 tgcctctgga tgtcgctcca caaggtaaac agttgattga actgcctgaa ctaccgcagc
     2221 cggagagcgc cgggcaactc tggctcacag tacgcgtagt gcaaccgaac gcgaccgcat
     2281 ggtcagaagc cgggcacatc agcgcctggc agcagtggcg tctggcggaa aacctcagtg
     2341 tgacgctccc cgccgcgtcc cacgccatcc cgcatctgac caccagcgaa atggattttt
     2401 gcatcgagct gggtaataag cgttggcaat ttaaccgcca gtcaggcttt ctttcacaga
     2461 tgtggattgg cgataaaaaa caactgctga cgccgctgcg cgatcagttc acccgtgcac
     2521 cgctggataa cgacattggc gtaagtgaag cgacccgcat tgaccctaac gcctgggtcg
     2581 aacgctggaa ggcggcgggc cattaccagg ccgaagcagc gttgttgcag tgcacggcag
     2641 atacacttgc tgatgcggtg ctgattacga ccgctcacgc gtggcagcat caggggaaaa
     2701 ccttatttat cagccggaaa acctaccgga ttgatggtag tggtcaaatg gcgattaccg
     2761 ttgatgttga agtggcgagc gatacaccgc atccggcgcg gattggcctg aactgccagc
     2821 tggcgcaggt agcagagcgg gtaaactggc tcggattagg gccgcaagaa aactatcccg
     2881 accgccttac tgccgcctgt tttgaccgct gggatctgcc attgtcagac atgtataccc
     2941 cgtacgtctt cccgagcgaa aacggtctgc gctgcgggac gcgcgaattg aattatggcc
     3001 cacaccagtg gcgcggcgac ttccagttca acatcagccg ctacagtcaa cagcaactga
     3061 tggaaaccag ccatcgccat ctgctgcacg cggaagaagg cacatggctg aatatcgacg
     3121 gtttccatat ggggattggt ggcgacgact cctggagccc gtcagtatcg gcggaattcc
     3181 agctgagcgc cggtcgctac cattaccagt tggtctggtg tcaaaaataa taataaccgg
     3241 gcaggccatg tctgcccgta tttcgcgtaa ggaaatccat tatgtactat ttaaaaaaca
     3301 caaacttttg gatgttcggt ttattctttt tcttttactt ttttatcatg ggagcctact
     3361 tcccgttttt cccgatttgg ctacatgaca tcaaccatat cagcaaaagt gatacgggta
     3421 ttatttttgc cgctatttct ctgttctcgc tattattcca accgctgttt ggtctgcttt
     3481 ctgacaaact cggaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat
     3541 cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact
     3601 catcaatgta tcttatcatg tctggatccc cgggggatcc actagttcta gactcgaagc
     3661 cgcggtgcgg gtgccagggc gtgcccttgg gctccccggg cgcgtactcc acctcaccca
     3721 tcgcggccgc caccgcggtg gagctccagc ttttgttccc tttagtgagg gttaattgcg
     3781 cgcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt
     3841 ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc
     3901 taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc
     3961 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct
     4021 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca
     4081 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
     4141 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt
     4201 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg
     4261 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc
     4321 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc
     4381 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc
     4441 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac
     4501 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt
     4561 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct
     4621 aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc
     4681 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt
     4741 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg
     4801 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc
     4861 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa
     4921 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag
     4981 gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg
     5041 tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga
     5101 gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag
     5161 cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa
     5221 gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc
     5281 atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca
     5341 aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg
     5401 atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat
     5461 aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc
     5521 aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg
     5581 gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg
     5641 gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt
     5701 gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca
     5761 ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata
     5821 ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac
     5881 atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa
     5941 gtgccaccta aattgtaagc gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa
     6001 tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat cccttataaa tcaaaagaat
     6061 agaccgagat agggttgagt gttgttccag tttggaacaa gagtccacta ttaaagaacg
     6121 tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac
     6181 catcacccta atcaagtttt ttggggtcga ggtgccgtaa agcactaaat cggaacccta
     6241 aagggagccc ccgatttaga gcttgacggg gaaagccggc gaacgtggcg agaaaggaag
     6301 ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg
     6361 taaccaccac acccgccgcg cttaatgcgc cgctacaggg cgcgtcccat tcgccattca
     6421 ggctgcgcaa ctgttgggaa gggcgatcgg tgcgggcctc ttcgctatta cgccagctgg
     6481 cgaaaggggg atgtgctgca aggcgattaa gttgggtaac gccagggttt tcccagtcac
     6541 gacgttgtaa aacgacggcc agtgagcgcg cgtaatacga ctcactatag ggcgaattgg
     6601 gtacatcgcg gccgcctcga agccgcggtg cgggtgccag ggcgtgccct tgggctcccc
     6661 gggcgcgtac tccacctcac ccatc
//
