LOCUS       sequence-313714-        4236 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pCR2.1-based plasmid holding a 309-bp fragment containing the 
            exon-1-exon-2 splice border of normal human beta-globin mRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4236)
  AUTHORS   Patsali P, Papasavva P, Christou S, Sitarou M, Antoniou MN, Lederer 
            CW, Kleanthous M
  TITLE     Relative and Absolute Quantification of Aberrant and Normal Splice 
            Variants in HBB(IVSI-110 (G > A)) beta-Thalassemia.
  JOURNAL   Int J Mol Sci. 2020 Sep 11;21(18). pii: ijms21186671. doi: 
            10.3390/ijms21186671.
  PUBMED    32933098
REFERENCE   2  (bases 1 to 4236)
  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: "Int J Mol 
            Sci."; date: "2020-09-11"; volume: "21(18). pii: ijms21186671. doi";
            pages: " 10.3390/ijms21186671"
COMMENT     Sequence Label: pCR2.1_HBB_N
FEATURES             Location/Qualifiers
     source          1..4236
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    101..122
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(137..142,143..160,161..167)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    175..191
                     /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)."
     primer_bind     180..202
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     199..215
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     199..215
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(663..682)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(664..682)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(689..706)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(689..705)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(698..720)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      846..1274
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(933..952)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1143..1164
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             1618..2412
                     /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(1672..1691)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2282..2301
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     CDS             join(2430..2498,2499..3290)
                     /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     complement(2648..2667)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3461..4049
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3950..3969
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4203..4220
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca
       61 ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtgagt tagctcactc
      121 attaggcacc ccaggcttta cactttatgc ttccggctcg tatgttgtgt ggaattgtga
      181 gcggataaca atttcacaca ggaaacagct atgaccatga ttacgccaag cttggtaccg
      241 agctcggatc cactagtaac ggccgccagt gtgctggaat tcggctttca ctagcaacct
      301 caaacagaca ccatggtgca cctgactcct gaggagaagt ctgccgttac tgccctgtgg
      361 ggcaaggtga acgtggatga agttggtggt gaggccctgg gcaggctgct ggtggtctac
      421 ccttggaccc agaggttctt tgagtccttt ggggatctgt ccactcctga tgctgttatg
      481 ggcaacccta aggtgaaggc tcatggcaag aaagtgctcg gtgcctttag tgatggcctg
      541 gctcacctgg acaacctcaa gggcaccttt gccacactga gtgagctgca ctgtgagccg
      601 aattctgcag atatccatca cactggcggc cgctcgagca tgcatctaga gggcccaatt
      661 cgccctatag tgagtcgtat tacaattcac tggccgtcgt tttacaacgt cgtgactggg
      721 aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca tccccctttc gccagctggc
      781 gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg
      841 aatggacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt
      901 gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct
      961 cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg
     1021 atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag
     1081 tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa
     1141 tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct attcttttga
     1201 tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa
     1261 atttaacgcg aattttaaca aaattcaggg cgcaagggct gctaaaggaa gcggaacacg
     1321 tagaaagcca gtccgcagaa acggtgctga ccccggatga atgtcagcta ctgggctatc
     1381 tggacaaggg aaaacgcaag cgcaaagaga aagcaggtag cttgcagtgg gcttacatgg
     1441 cgatagctag actgggcggt tttatggaca gcaagcgaac cggaattgcc agctggggcg
     1501 ccctctggta aggttgggaa gccctgcaaa gtaaactgga tggctttctt gccgccaagg
     1561 atctgatggc gcaggggatc aagatctgat caagagacag gatgaggatc gtttcgcatg
     1621 attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc
     1681 tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg
     1741 caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag
     1801 gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc
     1861 gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat
     1921 ctcctgtcat cccaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg
     1981 cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc
     2041 gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag
     2101 catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgcgcat gcccgacggc
     2161 gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc
     2221 cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata
     2281 gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc
     2341 gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac
     2401 gagttcttct gaattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta
     2461 ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag
     2521 taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca
     2581 gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta
     2641 aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc
     2701 gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc
     2761 ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca
     2821 ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc
     2881 acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca
     2941 taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac
     3001 tattaactgg cgaactactt actctagctt cccggcaaca attaatagac tggatggagg
     3061 cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg
     3121 ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg
     3181 gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac
     3241 gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc
     3301 aagtttactc atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct
     3361 aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc
     3421 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     3481 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     3541 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     3601 atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     3661 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     3721 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     3781 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     3841 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     3901 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     3961 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     4021 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     4081 tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg
     4141 ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc
     4201 gcagcgagtc agtgagcgag gaagcggaag agcgcc
//
