LOCUS       sequence-180623-        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"
     protein_bind    107..128
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(143..148,149..166,167..173)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    181..197
                     /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     186..208
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     205..221
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     205..221
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(689..708)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(690..708)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(715..732)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(715..731)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(724..746)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      872..1300
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(959..978)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1169..1190
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             1644..2438
                     /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(1698..1717)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2308..2327
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     CDS             join(2456..2524,2525..3316)
                     /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(2674..2693)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3487..4075
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3976..3995
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4229..4246
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
       61 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
      121 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
      181 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccaagcttg
      241 gtaccgagct cggatccact agtaacggcc gccagtgtgc tggaattcgg cttcacagtg
      301 cagctcactc agtgtggcaa aggtgccctt gaggttgtcc aggtgagcca ggccatcact
      361 aaaggcaccg agcactttct tgccatgagc cttcacctta gggttgccca taacagcatc
      421 aggagtggac agatccccaa aggactcaaa gaacctctgg gtccaagggt agaccaccag
      481 cagcctaagg gtgggaaaat agactgccca gggcctcacc accaacttca tccacgttca
      541 ccttgcccca cagggcagta acggcagact tctcctcagg agtcagatgc accatggtgt
      601 ctgtttgagg ttgctagtga aagccgaatt ctgcagatat ccatcacact ggcggccgct
      661 cgagcatgca tctagagggc ccaattcgcc ctatagtgag tcgtattaca attcactggc
      721 cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt acccaactta atcgccttgc
      781 agcacatccc cctttcgcca gctggcgtaa tagcgaagag gcccgcaccg atcgcccttc
      841 ccaacagttg cgcagcctga atggcgaatg gacgcgccct gtagcggcgc attaagcgcg
      901 gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct
      961 cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta
     1021 aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa
     1081 cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct
     1141 ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc
     1201 aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg
     1261 ttaaaaaatg agctgattta acaaaaattt aacgcgaatt ttaacaaaat tcagggcgca
     1321 agggctgcta aaggaagcgg aacacgtaga aagccagtcc gcagaaacgg tgctgacccc
     1381 ggatgaatgt cagctactgg gctatctgga caagggaaaa cgcaagcgca aagagaaagc
     1441 aggtagcttg cagtgggctt acatggcgat agctagactg ggcggtttta tggacagcaa
     1501 gcgaaccgga attgccagct ggggcgccct ctggtaaggt tgggaagccc tgcaaagtaa
     1561 actggatggc tttcttgccg ccaaggatct gatggcgcag gggatcaaga tctgatcaag
     1621 agacaggatg aggatcgttt cgcatgattg aacaagatgg attgcacgca ggttctccgg
     1681 ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg
     1741 atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc
     1801 tgtccggtgc cctgaatgaa ctgcaggacg aggcagcgcg gctatcgtgg ctggccacga
     1861 cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc
     1921 tattgggcga agtgccgggg caggatctcc tgtcatccca ccttgctcct gccgagaaag
     1981 tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat
     2041 tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg
     2101 tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca
     2161 ggctcaaggc gcgcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct
     2221 tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg
     2281 gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg
     2341 gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc
     2401 gcatcgcctt ctatcgcctt cttgacgagt tcttctgaat tgaaaaagga agagtatgag
     2461 tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt
     2521 tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt
     2581 gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga
     2641 acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat
     2701 tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga
     2761 gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag
     2821 tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg
     2881 accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg
     2941 ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt
     3001 agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg
     3061 gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc
     3121 ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg
     3181 tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac
     3241 ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact
     3301 gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa
     3361 acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa
     3421 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     3481 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     3541 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     3601 tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca
     3661 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     3721 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     3781 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
     3841 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
     3901 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
     3961 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
     4021 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
     4081 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt
     4141 tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac
     4201 cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaag
//
