LOCUS       sequence-313713-        4256 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pCR2.1-based plasmid holding a 328-bp fragment containing the 
            exon-1-exon-2 splice border of aberrant HBB[IVSI-110(G>A)] human 
            beta-globin mRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4256)
  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 4256)
  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_A
FEATURES             Location/Qualifiers
     source          1..4256
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     67..89
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     81..98
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     82..98
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     105..124
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        105..123
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(592..608)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(592..608)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(605..627)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    616..632
                     /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(640..646,647..664,665..670))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    685..706
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(823..840)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(994..1582)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1074..1093)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1753..2544,2545..2613))
                     /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     2376..2395
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     CDS             complement(2631..3425)
                     /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(2742..2761)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     3352..3371
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     rep_origin      complement(3769..4197)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3879..3900)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4091..4110
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 tattacgcca gctggcgaaa gggggatgtg ctgcaaggcg attaagttgg gtaacgccag
       61 ggttttccca gtcacgacgt tgtaaaacga cggccagtga attgtaatac gactcactat
      121 agggcgaatt gggccctcta gatgcatgct cgagcggccg ccagtgtgat ggatatctgc
      181 agaattcggc tttcactagc aacctcaaac agacaccatg gtgcacctga ctcctgagga
      241 gaagtctgcc gttactgccc tgtggggcaa ggtgaacgtg gatgaagttg gtggtgaggc
      301 cctgggcagt ctattttccc acccttaggc tgctggtggt ctacccttgg acccagaggt
      361 tctttgagtc ctttggggat ctgtccactc ctgatgctgt tatgggcaac cctaaggtga
      421 aggctcatgg caagaaagtg ctcggtgcct ttagtgatgg cctggctcac ctggacaacc
      481 tcaagggcac ctttgccaca ctgagtgagc tgcactgtga agccgaattc cagcacactg
      541 gcggccgtta ctagtggatc cgagctcggt accaagcttg gcgtaatcat ggtcatagct
      601 gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat
      661 aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc
      721 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
      781 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
      841 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
      901 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
      961 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     1021 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     1081 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     1141 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     1201 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     1261 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     1321 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     1381 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     1441 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     1501 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     1561 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     1621 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     1681 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     1741 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     1801 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     1861 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     1921 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     1981 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     2041 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     2101 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     2161 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     2221 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     2281 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     2341 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     2401 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     2461 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     2521 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     2581 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat tcagaagaac
     2641 tcgtcaagaa ggcgatagaa ggcgatgcgc tgcgaatcgg gagcggcgat accgtaaagc
     2701 acgaggaagc ggtcagccca ttcgccgcca agctcttcag caatatcacg ggtagccaac
     2761 gctatgtcct gatagcggtc cgccacaccc agccggccac agtcgatgaa tccagaaaag
     2821 cggccatttt ccaccatgat attcggcaag caggcatcgc catgggtcac gacgagatcc
     2881 tcgccgtcgg gcatgcgcgc cttgagcctg gcgaacagtt cggctggcgc gagcccctga
     2941 tgctcttcgt ccagatcatc ctgatcgaca agaccggctt ccatccgagt acgtgctcgc
     3001 tcgatgcgat gtttcgcttg gtggtcgaat gggcaggtag ccggatcaag cgtatgcagc
     3061 cgccgcattg catcagccat gatggatact ttctcggcag gagcaaggtg ggatgacagg
     3121 agatcctgcc ccggcacttc gcccaatagc agccagtccc ttcccgcttc agtgacaacg
     3181 tcgagcacag ctgcgcaagg aacgcccgtc gtggccagcc acgatagccg cgctgcctcg
     3241 tcctgcagtt cattcagggc accggacagg tcggtcttga caaaaagaac cgggcgcccc
     3301 tgcgctgaca gccggaacac ggcggcatca gagcagccga ttgtctgttg tgcccagtca
     3361 tagccgaata gcctctccac ccaagcggcc ggagaacctg cgtgcaatcc atcttgttca
     3421 atcatgcgaa acgatcctca tcctgtctct tgatcagatc ttgatcccct gcgccatcag
     3481 atccttggcg gcaagaaagc catccagttt actttgcagg gcttcccaac cttaccagag
     3541 ggcgccccag ctggcaattc cggttcgctt gctgtccata aaaccgccca gtctagctat
     3601 cgccatgtaa gcccactgca agctacctgc tttctctttg cgcttgcgtt ttcccttgtc
     3661 cagatagccc agtagctgac attcatccgg ggtcagcacc gtttctgcgg actggctttc
     3721 tacgtgttcc gcttccttta gcagcccttg cgccctgaat tttgttaaaa ttcgcgttaa
     3781 atttttgtta aatcagctca ttttttaacc aataggccga aatcggcaaa atcccttata
     3841 aatcaaaaga atagaccgag atagggttga gtgttgttcc agtttggaac aagagtccac
     3901 tattaaagaa cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc
     3961 cactacgtga accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa
     4021 atcggaaccc taaagggagc ccccgattta gagcttgacg gggaaagccg gcgaacgtgg
     4081 cgagaaagga agggaagaaa gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg
     4141 tcacgctgcg cgtaaccacc acacccgccg cgcttaatgc gccgctacag ggcgcgtcca
     4201 ttcgccattc aggctgcgca actgttggga agggcgatcg gtgcgggcct cttcgc
//
