Supplementary Data
Table of Contents
Supplementary Materials and Methods
1.Vector construction
2.Subcloning of target genes to the entry vectors
3.Pimers used in this report
Supplementary Data for Vector Information
Vector information for pL12R34-CmR-ccdB
Vector information for pL34R12-CmR-ccdB
Vector information for pL12R34-Ap
Vector information for pL34R12-Cm
Vector information for pL12R34-sacB
Vector information for pL34R12-ccdB
Vector information for pL12R34H-Ap
Vector information for pL34R12H-Cm
Vector information for pL12R34H-sacB
Vector information for pL34R12H-CmR-ccdB
Vector information for pGWTAC
Note: The vector sequences have been compiled from information in the sequence databases, published literature, and other sources, together with partial sequences obtained by us. These vectors have not been completely sequenced.All of the vectors described in this report will be made freely available for noncommercialresearch purposes.
Supplementary Materials and Methods
- Vector construction
Two parental entry vectors, pL12R34-CmR-ccdB and pL34R12-CmR-ccdB were constructed. The pL12R34-CmR-ccdB vector was derived sequentially from the intermediate vectors pGL12-ApR, pGL12, pGEN, pGL12P and pENTR1A (Invitrogen, Carlsbad, CA) and the pL34R12-CmR-ccdB vector was derived sequentially from the intermediate vectors pGL34-ApR, pGL34-CmR, pGEN-ApR and pGL12. All of the ccdB-containing vectors were maintained and propagated in E.coli strain DB3.1 (Invitrogen), and all of PCR products were amplified with proofreading Pfu DNA polymerase and verified by sequencing when necessary.
To construct pGL12P, 1AF and 1AR primers (Table S1) were used to amplify sequences from the pENTR1A vector, and GenF and GenR primers (Table S1) were used for PCR amplification from the pEZ-TNL vector(kindly provided by Dr. DavidW.Ehrhardt, Carnegie Institution of Washington, Stanford, CA). The former PCR products were digested with BspHI, and the latter PCR products were double-digested with BspHI and SmaI. Then the two PCR fragments were ligated to generate pGL12P.
To construct pGEN, three pairs of primers (Table S1) were used sequentially for PCR amplification. EGFPF1 and EGFPR1 primers (Table S1) were used for the first PCR amplification from pEZ-TNL. The PCR products were used as templates for the second PCR amplification with EGFPF2 and EGFPR2 as primers (Table S1). Then the third PCR amplification was carried out with the second PCR products as templates and with EGFPF3 and EGFPR3 as primers (Table S1). The PCR products were double-digested with AflII and BspHI and inserted into the AfllI and BspHI sites in pGL12P to generate pGEN. The small ApaI-PstI fragment from pENTR1A was inserted into the ApaI and PstI sites of pGEN to generate the pGL12.
To construct pGL12-ApR, ApRF1 and ApRR1 primers (Table S1) were used for PCR amplification from pUC18. The PCR fragments were digested with NheI and inserted into the DraI and XbaI sites in pGL12, the resultant vector was named pGL12-ApR. To construct pL12R34-CmR-ccdB, R34F and R34R primers (Table S1) were used for PCR amplification from pDESTR4R3 (Invitrogen). The PCR products were digested with AscI and PacI and inserted into the AscI and PacI sites of pGL12-ApR to generate pL12R34-CmR-ccdB.
To obtain attL3-attL4 cassettes, CmR-B4F/CmR-B1R primer pairs and CmR-B2F/CmR-B3R primer pairs (Table S1) were used for PCR amplification from pCAMBIA1200, respectively. Then BP recombination reactions were carried out between the CmR-B4F/CmR-B1R PCR fragments and pDONRP4P1R (Invitrogen) and between the CmR-B2F/CmR-B3R PCR fragments and pDONRP2RP3 (Invitrogen). PCRs and in vitro BP recombination reactions were carried out according to the manufacturer’s instructions (Invitrogen). The resultant entry vectors were named pL4R1-CmR and pL3R2-CmR. The small AflII-HindIII fragment from pL4R1-CmR was inserted into the AflII and HindIII sites of pL3R2-CmR to generate pKL34. The small SacI fragment of pKL34 was polished with T4 DNA polymerase, an A residue was added to its 3’ end with Taq polymerase and the entire sequence was cloned into the pMD18-T vector (TaKaRa) to generate pAL34-CmR.
To construct pGEN-ApR, ApRF2 and ApRR2 primers (Table S1) were used for PCR amplification from pUC18. The PCR fragments were digested with ApaI and PstI, and inserted into the ApaI and PstI sites of pGL12, the resultant vector was named pGEN-ApR. To construct pGL34-CmR, the small XbaI-SalI fragment from pAL34-CmR was inserted into SpeI and XhoI sites of pGEN-ApR to generate pGL34-CmR. The small HindIII-PacI fragment of pGL12-ApR was inserted into the HindIII and PacI sites of pGL34-CmR to generate pGL34-ApR. The small AscI-PacI fragment of pMDC32 (kindly provided by Dr.MarkD.Curtis, University of Zurich, Zurich, Switzerland)was inserted into pGL34-ApR to generatepL34R12-CmR-ccdB.
To construct pL12R34-sacB (pL12R34sB), the large NotI-SalI fragment of pL12R34-CmR-ccdB was polished with T4 DNA polymerase and dephosphorylated with Antarctic Phosphatase (NEB). Then the fragment was ligated with the small AatII fragment of pDNR-Dual (BD Biosciences Clontech) to generate pL14sB. To construct pL34R12-ccdB (pL34R12cB), the large NotI-BamHI fragment of pL34R12-CmR-ccdB was polished and self-ligated to generate pL34R12cB.
To construct pL12R34-Ap (pL12R34A), ApRF3 and ApRR3 primers (Table S1) were used for PCR amplification from pUC18. The PCR products were double-digested with EagI and XhoI and inserted into the NotI and SalI sites of the pL12R34-CmR-ccdB to generate pL12R34A. To construct pL34R12-Cm (pL34R12C), CmRF1 and CmRR1 primers (Table S1) were used for PCR amplification from pCAMBIA1200. The PCR products were double-digested with EagI and XhoI and inserted into the NotI and SalI sites of pL34R12-CmR-ccdB to generate pL34R12C.
To construct improved versions of the vector system, I-SceI and PI-SceI homing endonuclease sites were introduced into the vector system. To introduce the PI-SceI sites, PI-CmRF and PI-CmRR primers (Table S1) were used for PCR amplification from pCAMBIA1200, and the PCR products were used as templates for the second round of PCR with single PI-FR as primer (Table S1). pL12R34-CmR-ccdB and pL34R12-CmR-ccdB were digested with NotI and SalI, respectively, and polished with T4 DNA polymerase; the fragments were ligated with the PCR products to generate pL14R32-PI-CmR and pL34R12-PI-CmR, respectively. Similarly, PI-ApRF and PI-ApRR primers (Table S1) were used for PCR amplification from pUC18, and then the single PI-FR primer (Table S1) was used for the second PCR amplification. The PCR products were inserted into pL12R34-CmR-ccdB to generate pL14R32-PI-ApR. To introduce the I-SceI sites, the two oligonucleotides, I-BPF and I-BPR, were annealed to form a linker. The linker was inserted into the BspHI and PstI sites of pL14R32-PI-CmR and pL34R12-PI-CmR to generate pL12R34-IPI-C and pL34R12-IPI-C, respectively. Then, another linker formed by two oligonucleotides, I-AAF and I-AAR, was inserted into the AflII and ApaI sites of pL12R4-IPI-C and pL34R12-IPI-C to generate pL12R34HC and pL34R12HC, respectively. The small PI-SceI fragment of pL12R34-PI-A was inserted into the PI-SceI sites of pL12R34HC to generate pL12R34HA.
To construct pL12R34HsB, sacBF and sacBR primes were used for PCR amplification from the pDNR-Dual. The PCR products were digested with BamHI and XhoI and inserted into the BamHI and SalI sites of pL12R34-CmR-ccdB to generate pL12R34sB2. Then a linker formed by PI-BNF and PI-BNR was inserted into the pL12R34sB2 to generate pL12R34PIsB. The small AscI-PacI fragment of pL12R34PIsB was inserted into the AscI and PacI sites of the pL12R34HC to generate pL12R34HsB.
To construct pL34R12HCcB, the pL34R12-CmR-ccdB was double-digested with NotI and BamHI and polished with T4 DNA polymerase. Then, the fragment was ligated with the small PI-SceI fragment of pL12R34HC to generate pL34R12PICcB. The small AscI-PacI fragment of pL34R12PICcB was inserted into the AscI and PacI sites of pL34R12HC to generate pL34R12HCcB.
To construct a Gateway-compatible TAC vector, pGWTAC, 15F and 15R primers (Table S1) were used for PCR amplification from pDEST15 (Invitrogen) in which the NotI site had been disrupted. The PCR products were digested with HindIII and NotI and inserted into the HindIII and NotI sites of pYLTAC747 (kindly provided by Professor Yao-Guang Liu,South China Agricultural University, Guangzhou, China) to generate pGWTAC.
- Subcloning of target genes to the entry vectors
To construct pL12R34sB-MAR, and pL34R12cB-MAR, CmRF2 and CmRR2 primers (Table S1) were used for PCR amplification from pCAMBIA1200, digested with PacI and PvuI and inserted into the PacI sites of pL12R34sB and pL34R12cB to generate pL12R34sB-CmR and pL34R12cB-CmR, respectively. The first half of TM1 (RB7 MAR element) was amplified using TM1-1F and TM1-1R primers (Table S1), digested with PacI and inserted into the PacI and PmeI sites of pL34R12cB-CmR to generate pL34R12cB-TM1-1; the second half of TM1 was amplified using TM1-2F and TM1-2R primers (Table S1), digested with PacI and inserted into the PacI and PmeI sites of pL12R34sB-CmR to generate pL12R34sB-TM1-2. TM2F and TM2R were used for PCR amplification of another MAR sequence–TM2 (kindly provided by ProfessorCheng-ChaoZheng,ShandongAgriculturalUniversity, Jinan, China), the PCR products were digested with HindIII and EcoRI and inserted into the HindIII and EcoRI sites of pL12R34sB-TM1-2 and pL34R12cB-TM1-1 to generate pL12R34sB-MAR and pL34R12cB-MAR, respectively. The large AscI-PacI fragment of pL34R12cB-MAR was polished and self-ligated to generate pL34-MAR. The small HindIII-EcoRI fragment of pBI121 was inserted into the HindIII and EcoRI sites of pL34R12cB-TM1-1 and pL12R34sB-TM1-2 to generate pL12R34sB-GUS and pL34R12cB-GUS, respectively. To construct pL12R34sB-EGFP, GFPF and GFPR primers (Table S1) were used for PCR amplification from the pEZ-TNL, the PCR products were digested with XbaI and SacI and inserted into the XbaI and SacI sites of pL12R34sB-GUS to generate pL12R34sB-EGFP. The Small HindIII-EcoRI fragments of pL34R12cB-MAR and pL12R34sB-EGFP were inserted into the HindIII and EcoRI sites of the pL12R34A to generate pL12R34A-MAR and pL12R34A-EGFP, respectively. The Small HindIII-EcoRI fragments of pL34R12cB-MAR and pL34R12cB-GUS were inserted into the HindIII and EcoRI sites of the pL34R12C to generate pL34R12C-MAR and pL34R12C-EGFP, respectively.
- Pimers used in this report
Table S1 Primers used in this report
Primer names / Sequences (5'→3')1AF / GCCCCGTTCCACTGAGCGTCAGACCCGGTAG
1AR / TGCGTCATGAGAGATTTTGAGACACGGGCCAGAG
GenF / AAAATCATGAGACGCACACCGTGGAAACGGATGAAGGCAC
GenR / AAAACCCGGGCGGCGTTGTGACAATTTACCGAACAACTCC
EGFPF1 / CATACGATTTAGGTGACACTATAGAAGGGCCCACAAGTTCAGCGTGTCCG
EGFPR1 / GTAATACGACTCACTATAGGGCCTGCAGTTCACCTTGATGCCGTTC
EGFPF2 / ACGACGTTGTAAAACGACGGCCAGTGCATACGATTTAGGTGACACTATAG
EGFPR2 / ACAGGAAACAGCTATGACCATGATTACGGTAATACGACTCACTATAGGGC
EGFPF3 / AAAACTTAAGCGCCAGGGTTTTCCCAGTCACGACGTTGTAAAACGACGGCC
EGFPR3 / AGCCTCATGAGAGCGGATAACAATTTCACACAGGAAACAGCTATGACCATG
ApRF1 / ACTGAAGCTTCGGAATTCGGCGCGCCCTTTTCGGGGAAATGTG
ApRR1 / ACTGGCTAGCTTAATTAAGGCGCGCCTGACGCTCAGTGGAACG
R34F / GCGGTTAATTAACATGATTACGCCAAGCTATC
R34R / TATTTAGGCGCGCCAGTGAATTATCAACTATG
ApRF2 / ATCGGGGCCCACTAGTTTCGGGGAAATGTGCGC
ApRR2 / ATGCCTGCAGCTCGAGTGACGCTCAGTGGAACG
CmR-B4F / GGGGACAACTTTGTATAGAAAAGTTGTTAATTAAGTGGACATAAGCCTCGTTC
CmR-B1R / GGGGACTGCTTTTTTGTACAAACTTGAAGCTTGACTTGACCTGATAGTTTGGC
CmR-B2F / GGGGACAGCTTTCTTGTACAAAGTGGTTAATTAAGTGGACATAAGCCTCGTTC
CmR-B3R / GGGGACAACTTTGTATAATAAAGTTGAAGCTTGACTTGACCTGATAGTTTGGC
ApRF3 / AACTCGGCCGTTCGGGGAAATGTGCGC
ApRR3 / ACGGCTCGAGTGACGCTCAGTGGAACG
CmRF1 / AATTCGGCCGGTGGACATAAGCCTCGTTCGGTTCG
CmRR1 / GCGGCTCGAGGACTTGACCTGATAGTTTGGCTGTG
PI-CmRF / AGAGGTAATGAAATGGCAACATAAGCCTCGTTCGGTTC
PI-CmRR / AGAGGTAATGAAATGGCAGACCTGATAGTTTGGCTGTG
PI-FR / ATCTATGTCGGGTGCGGAGAAAGAGGTAATGAAATGGC
PI-ApRF / AGAGGTAATGAAATGGCATTCGGGGAAATGTGCGC
PI-ApRR / AGAGGTAATGAAATGGCATGACGCTCAGTGGAACG
I-BPF / GATTACCCTGTTATCCCTAT
I-BPR / CATGATAGGGATAACAGGGTAATCTGCA
I-AAF / TTAAGTAGGGATAACAGGGTAATGGGCC
I-AAR / CATTACCCTGTTATCCCTAC
PI-BNF / GGCCCGTGCCATTTCATTACCTCTTTCTCCGCACCCGACATAGATC
PI-BNR / GATCGATCTATGTCGGGTGCGGAGAAAGAGGTAATGAAATGGCACG
15F / GGCCCCAAGCTTCGACCATCCTCCAAAATCGGATCTG
15R / AATTGCGGCCGCTCCTTTCGGGCTTTGTTAGCAGCCG
CmRF2 / CGCGTTAATTAACCAGTTTAAACGTGGACATAAGCCTCGTTC
CmRR2 / AAATTCGCGATCGGTTTAAACGACTTGACCTGATAGTTTGGC
TM1-1F / CGCGTTAATTAAAAATCCCAATTATATTTGGTC
TM1-1R / AAACGTTTTGCCGGGTTTTCAGCATTTTC
TM1-2F / AGCCTTAATTAACCGAACCAATCCAAACCG
TM1-2R / AAACGCAGAAGAAGTTCCCAATAGTAGTCC
TM2F / ACACAAGCTTCCCAATTATTGCGAGTGAAAGCTGCTC
TM2R / GCGTGAATTCCCAATCTTGAAATATCGAAGGTTCGCC
GFPF / ATATCTAGAATGGTGAGCAAGGGCG
GFPR / ATGGAGCTCTTACTTGTACAGCTCG
The restriction enzyme sites are indicated by the boxes.
Supplementary Data for Vector Information
Vector information for pL12R34-CmR-ccdB
Map of pL12R34-CmR-ccdB
Location of Features
•ccdB: 253-558
•attR4: 598-722 (c)
•attL2: 755-854 (c)
•T7: 870-889 (c)
•RV-M: 914-937 (c)
•Gen(R): 1199-1732
•pUCorigin: 1825-2498
•rrnB T1: 2607-2650 (c)
•rrnB T2: 2782-2825 (c)
•M13-47: 2850-2873
•SP6: 2895-2918
•attL1: 2927-3026
•attR3: 3065-3189
•Cm(R): 3298-3957
(c) = complementary strand
Sequence of pL12R34-CmR-ccdB
tacccgaagtatgtcaaaaagaggtatgctatgaagcagcgtattacagtgacagttgacagcgacagctatcagttgctcaaggcatatatgatgtcaatatctccggttcggtaagcacaaccatgcagaatgaagcccgtcgtctgcgtgccgaacgctggaaagcggaaaatcaggaagggatggctgaggtcgcccggtttattgaaatgaacggctcttttgccgacgagaacaggggctggtgaaatgcagtttaaggtttacacctataaaagagagagccgttatcgtctgtttgtggatgtacagagtgatattattgacacgcccgggcgacggatggtgatccccctggccagtgcacgtctgctgtcagataaagtcccccgtgaactttacccggtggtgcatatcggggatgaaagctggcgcatgatgaccaccgatatggccagtgtgccggtctccgttatcggggaagaagtggctgatctcagccaccgcgaaaatgacatcaaaaacgccattaacctgatgttctggggaatataaatgtcaggctcccttatacacagccagtctgcaggtcgaccatagtgactggatatgttgtgttttacagtattatgtagtctgttttttatgcaaaatctaatttaatatattgatatttatatcattttacgtttctcgttcaacttttctatacaaagttgatagcttggcgtaatcatgttaattaagctagacccagctttcttgtacaaagttggcattataagaaagcattgcttatcaatttgttgcaacgaacaggtcactatcagtcaaaataaaatcattatttgccatccagctgcaggccctatagtgagtcgtattaccgtaatcatggtcatagctgtttcctgtgtgaaattgttatccgctctcatgagacgcacaccgtggaaacggatgaaggcacgaacccagttgacataagcctgttcggttcgtaaactgtaatgcaagtagcgtatgcgctcacgcaactggtccagaaccttgaccgaacgcagcggtggtaacggcgcagtggcggttttcatggcttgttatgactgtttttttgtacagtctatgcctcgggcatccaagcagcaagcgcgttacgccgtgggtcgatgtttgatgttatggagcagcaacgatgttacgcagcagcaacgatgttacgcagcagggcagtcgccctaaaacaaagttaggtggctcaagtatgggcatcattcgcacatgtaggctcggccctgaccaagtcaaatccatgcgggctgctcttgatcttttcggtcgtgagttcggagacgtagccacctactcccaacatcagccggactccgattacctcgggaacttgctccgtagtaagacattcatcgcgcttgctgccttcgaccaagaagcggttgttggcgctctcgcggcttacgttctgcccaggtttgagcagccgcgtagtgagatctatatctatgatctcgcagtctccggcgagcaccggaggcagggcattgccaccgcgctcatcaatctcctcaagcatgaggccaacgcgcttggtgcttatgtgatctacgtgcaagcagattacggtgacgatcccgcagtggctctctatacaaagttgggcatacgggaagaagtgatgcactttgatatcgacccaagtaccgccacctaacaattcgttcaagccgagatcggcttcccggccgcggagttgttcggtaaattgtcacaacgccgcccgccccgttccactgagcgtcagacccggtagaaaagatcaaaggatcttcttgagatcctttttttctgcgcgtaatctgctgcttgcaaacaaaaaaaccaccgctaccagcggtggtttgtttgccggatcaagagctaccaactctttttccgaaggtaactggcttcagcagagcgcagataccaaatactgttcttctagtgtagccgtagttaggccaccacttcaagaactctgtagcaccgcctacatacctcgctctgctaatcctgttaccagtggctgctgccagtggcgataagtcgtgtcttaccgggttggactcaagacgatagttaccggataaggcgcagcggtcgggctgaacggggggttcgtgcacacagcccagcttggagcgaacgacctacaccgaactgagatacctacagcgtgagctatgagaaagcgccacgcttcccgaagggagaaaggcggacaggtatccggtaagcggcagggtcggaacaggagagcgcacgagggagcttccagggggaaacgcctggtatctttatagtcctgtcgggtttcgccacctctgacttgagcgtcgatttttgtgatgctcgtcaggggggcggagcctatggaaaaacgccagcaacgcggcctttttacggttcctggccttttgctggccttttgctcacatgttctttcctgcgttatcccctgattctgtggataaccgtattaccgctagcatggatctcggggacgtctaactactaagcgagagtagggaactgccaggcatcaaataaaacgaaaggctcagtcggaagactgggcctttcgttttatctgttgtttgtcggtgaacgctctcctgagtaggacaaatccgccgggagcggatttgaacgttgtgaagcaacggcccggagggtggcgggcaggacgcccgccataaactgccaggcatcaaactaagcagaaggccatcctgacggatggcctttttgcgtttctacaaactcttcctgttagttagttacttaagcgccagggttttcccagtcacgacgttgtaaaacgacggccagtgcatacgatttaggtgacactatagaagggccccaaataatgattttattttgactgatagtgacctgttcgttgcaacaaattgataagcaatgcttttttataatgccaactttgtacaaaaaagcaggctttactgaagcttcggaattcggcgcgccagtgaattatcaactatgtataataaagttgaacgagaaacgtaaaatgatataaatatcaatatattaaattagattttgcataaaaaacagactacataatactgtaaaacacaacatatccagtcactatggcggccgcattaggcacccccaggctttacactttatgcttccggctcgtataatgtgtggattttgagttaggatccgtcgagattttcaggagctaaggaagctaaaatggagaaaaaaatcactggatataccaccgttgatatatcccaatggcatcgtaaagaacattttgaggcatttcagtcagttgctcaatgtacctataaccagaccgttcagctggatattacggcctttttaaagaccgtaaagaaaaataagcacaagttttatccggcctttattcacattcttgcccgcctgatgaatgctcatccggaattccgtatggcaatgaaagacggtgagctggtgatatgggatagtgttcacccttgttacaccgttttccatgagcaaactgaaacgttttcatcgctctggagtgaataccacgacgatttccggcagtttctacacatatattcgcaagatgtggcgtgttacggtgaaaacctggcctatttccctaaagggtttattgagaatatgtttttcgtctcagccaatccctgggtgagtttcaccagttttgatttaaacgtggccaatatggacaacttcttcgcccccgttttcaccatgggcaaatattatacgcaaggcgacaaggtgctgatgccgctggcgattcaggttcatcatgccgtttgtgatggcttccatgtcggcagaatgcttaatgaattacaacagtactgcgatgagtggcagggcggggcgtaatctagaggatccggcttactaaaagccagataacagtatgcgtatttgcgcgctgatttttgcggtataagaatatatactgatatgta
Vector information for pL34R12-CmR-ccdB
Map of pL34R12-CmR-ccdB
Location of Features
•ccdB: 253-558
•attR2: 599-723 (c)
•attL4: 743-838 (c)
•T7: 858-877
•RV-M: 902-925
•Gen(R): 1187-1720
•pUCorigin: 1813-2486
•rrnB T1: 2595-2638 (c)
•rrnB T2: 2770-2813 (c)
•M13-47: 2838-2861 (c)
•SP6: 2883-2906 (c)
•attL3: 2927-3022
•attR1: 3055-3179
•Cm(R): 3288-3947
(c) = complementary strand
Sequence of pL34R12-CmR-ccdB
tacccgaagtatgtcaaaaagaggtgtgctatgaagcagcgtattacagtgacagttgacagcgacagctatcagttgctcaaggcatatatgatgtcaatatctccggtctggtaagcacaaccatgcagaatgaagcccgtcgtctgcgtgccgaacgctggaaagcggaaaatcaggaagggatggctgaggtcgcccggtttattgaaatgaacggctcttttgctgacgagaacagggactggtgaaatgcagtttaaggtttacacctataaaagagagagccgttatcgtctgtttgtggatgtacagagtgatattattgacacgcccgggcgacggatggtgatccccctggccagtgcacgtctgctgtcagataaagtctcccgtgaactttacccggtggtgcatatcggggatgaaagctggcgcatgatgaccaccgatatggccagtgtgccggtctccgttatcggggaagaagtggctgatctcagccaccgcgaaaatgacatcaaaaacgccattaacctgatgttctggggaatataaatgtcaggctcccttatacacagccagtctgcaggtcgaccatagtgactggatatgttgtgttttacagtattatgtagtctgttttttatgcaaaatctaatttaatatattgatatttatatcattttacgtttctcgttcagctttcttgtacaaagtggttcgataattccttaattaacaacttttctatacaaagttggcattataaaaaagcattgcttatcaatttgttgcaacgaacaggtcactatcagtcaaaataaaatcattatttggaatcgtcgagctgcaggccctatagtgagtcgtattaccgtaatcatggtcatagctgtttcctgtgtgaaattgttatccgctctcatgagacgcacaccgtggaaacggatgaaggcacgaacccagttgacataagcctgttcggttcgtaaactgtaatgcaagtagcgtatgcgctcacgcaactggtccagaaccttgaccgaacgcagcggtggtaacggcgcagtggcggttttcatggcttgttatgactgtttttttgtacagtctatgcctcgggcatccaagcagcaagcgcgttacgccgtgggtcgatgtttgatgttatggagcagcaacgatgttacgcagcagcaacgatgttacgcagcagggcagtcgccctaaaacaaagttaggtggctcaagtatgggcatcattcgcacatgtaggctcggccctgaccaagtcaaatccatgcgggctgctcttgatcttttcggtcgtgagttcggagacgtagccacctactcccaacatcagccggactccgattacctcgggaacttgctccgtagtaagacattcatcgcgcttgctgccttcgaccaagaagcggttgttggcgctctcgcggcttacgttctgcccaggtttgagcagccgcgtagtgagatctatatctatgatctcgcagtctccggcgagcaccggaggcagggcattgccaccgcgctcatcaatctcctcaagcatgaggccaacgcgcttggtgcttatgtgatctacgtgcaagcagattacggtgacgatcccgcagtggctctctatacaaagttgggcatacgggaagaagtgatgcactttgatatcgacccaagtaccgccacctaacaattcgttcaagccgagatcggcttcccggccgcggagttgttcggtaaattgtcacaacgccgcccgccccgttccactgagcgtcagacccggtagaaaagatcaaaggatcttcttgagatcctttttttctgcgcgtaatctgctgcttgcaaacaaaaaaaccaccgctaccagcggtggtttgtttgccggatcaagagctaccaactctttttccgaaggtaactggcttcagcagagcgcagataccaaatactgttcttctagtgtagccgtagttaggccaccacttcaagaactctgtagcaccgcctacatacctcgctctgctaatcctgttaccagtggctgctgccagtggcgataagtcgtgtcttaccgggttggactcaagacgatagttaccggataaggcgcagcggtcgggctgaacggggggttcgtgcacacagcccagcttggagcgaacgacctacaccgaactgagatacctacagcgtgagctatgagaaagcgccacgcttcccgaagggagaaaggcggacaggtatccggtaagcggcagggtcggaacaggagagcgcacgagggagcttccagggggaaacgcctggtatctttatagtcctgtcgggtttcgccacctctgacttgagcgtcgatttttgtgatgctcgtcaggggggcggagcctatggaaaaacgccagcaacgcggcctttttacggttcctggccttttgctggccttttgctcacatgttctttcctgcgttatcccctgattctgtggataaccgtattaccgctagcatggatctcggggacgtctaactactaagcgagagtagggaactgccaggcatcaaataaaacgaaaggctcagtcggaagactgggcctttcgttttatctgttgtttgtcggtgaacgctctcctgagtaggacaaatccgccgggagcggatttgaacgttgtgaagcaacggcccggagggtggcgggcaggacgcccgccataaactgccaggcatcaaactaagcagaaggccatcctgacggatggcctttttgcgtttctacaaactcttcctgttagttagttacttaagcgccagggttttcccagtcacgacgttgtaaaacgacggccagtgcatacgatttaggtgacactatagaagggcccactagagattccaaataatgattttattttgactgatagtgacctgttcgttgcaacaaattgataagcaatgcttttttataatgccaactttgtataataaagttgaagcttcggaattcggcgcgccaagctatcaaacaagtttgtacaaaaaagctgaacgagaaacgtaaaatgatataaatatcaatatattaaattagattttgcataaaaaacagactacataatactgtaaaacacaacatatccagtcactatggcggccgcattaggcaccccaggctttacactttatgcttccggctcgtataatgtgtggattttgagttaggatccggcgagattttcaggagctaaggaagctaaaatggagaaaaaaatcactggatataccaccgttgatatatcccaatggcatcgtaaagaacattttgaggcatttcagtcagttgctcaatgtacctataaccagaccgttcagctggatattacggcctttttaaagaccgtaaagaaaaataagcacaagttttatccggcctttattcacattcttgcccgcctgatgaatgctcatccggaattccgtatggcaatgaaagacggtgagctggtgatatgggatagtgttcacccttgttacaccgttttccatgagcaaactgaaacgttttcatcgctctggagtgaataccacgacgatttccggcagtttctacacatatattcgcaagatgtggcgtgttacggtgaaaacctggcctatttccctaaagggtttattgagaatatgtttttcgtctcagccaatccctgggtgagtttcaccagttttgatttaaacgtggccaatatggacaacttcttcgcccccgttttcaccatgggcaaatattatacgcaaggcgacaaggtgctgatgccgctggcgattcaggttcatcatgccgtctgtgatggcttccatgtcggcagaatgcttaatgaattacaacagtactgcgatgagtggcagggcggggcgtaatctagaggatccggcttactaaaagccagataacagtatgcgtatttgcgcgctgatttttgcggtataagaatatatactgatatgta
Vector information for pL12R34-Ap
Map of pL12R34-Ap
Location of Features
•Ap(R): 3287-423
•attR4: 570-694 (c)
•attL2: 727-826 (c)
•T7: 842-861 (c)
•RV-M: 886-909 (c)
•Gen(R): 1171-1704
•pUCorigin: 1797-2470
•rrnB T1: 2579-2622 (c)
•rrnB T2: 2754-2797 (c)
•M13-47: 2822-2845
•SP6: 2867-2890
•attL1: 2899-2998
•attR3: 3037-3161
(c) = complementary strand
Sequence of pL12R34-Ap
accgcttttttgcacaacatgggggatcatgtaactcgccttgatcgttgggaaccggagctgaatgaagccataccaaacgacgagcgtgacaccacgatgcctgtagcaatggcaacaacgttgcgcaaactattaactggcgaactacttactctagcttcccggcaacaattaatagactggatggaggcggataaagttgcaggaccacttctgcgctcggcccttccggctggctggtttattgctgataaatctggagccggtgagcgtgggtctcgcggtatcattgcagcactggggccagatggtaagccctcccgtatcgtagttatctacacgacggggagtcaggcaactatggatgaacgaaatagacagatcgctgagataggtgcctcactgattaagcattggtaactgtcagaccaagtttactcatatatactttagattgatttaaaacttcatttttaatttaaaaggatctaggtgaagatcctttttgataatctcatgaccaaaatcccttaacgtgagttttcgttccactgagcgtcactcgaccatagtgactggatatgttgtgttttacagtattatgtagtctgttttttatgcaaaatctaatttaatatattgatatttatatcattttacgtttctcgttcaacttttctatacaaagttgatagcttggcgtaatcatgttaattaagctagacccagctttcttgtacaaagttggcattataagaaagcattgcttatcaatttgttgcaacgaacaggtcactatcagtcaaaataaaatcattatttgccatccagctgcaggccctatagtgagtcgtattaccgtaatcatggtcatagctgtttcctgtgtgaaattgttatccgctctcatgagacgcacaccgtggaaacggatgaaggcacgaacccagttgacataagcctgttcggttcgtaaactgtaatgcaagtagcgtatgcgctcacgcaactggtccagaaccttgaccgaacgcagcggtggtaacggcgcagtggcggttttcatggcttgttatgactgtttttttgtacagtctatgcctcgggcatccaagcagcaagcgcgttacgccgtgggtcgatgtttgatgttatggagcagcaacgatgttacgcagcagcaacgatgttacgcagcagggcagtcgccctaaaacaaagttaggtggctcaagtatgggcatcattcgcacatgtaggctcggccctgaccaagtcaaatccatgcgggctgctcttgatcttttcggtcgtgagttcggagacgtagccacctactcccaacatcagccggactccgattacctcgggaacttgctccgtagtaagacattcatcgcgcttgctgccttcgaccaagaagcggttgttggcgctctcgcggcttacgttctgcccaggtttgagcagccgcgtagtgagatctatatctatgatctcgcagtctccggcgagcaccggaggcagggcattgccaccgcgctcatcaatctcctcaagcatgaggccaacgcgcttggtgcttatgtgatctacgtgcaagcagattacggtgacgatcccgcagtggctctctatacaaagttgggcatacgggaagaagtgatgcactttgatatcgacccaagtaccgccacctaacaattcgttcaagccgagatcggcttcccggccgcggagttgttcggtaaattgtcacaacgccgcccgccccgttccactgagcgtcagacccggtagaaaagatcaaaggatcttcttgagatcctttttttctgcgcgtaatctgctgcttgcaaacaaaaaaaccaccgctaccagcggtggtttgtttgccggatcaagagctaccaactctttttccgaaggtaactggcttcagcagagcgcagataccaaatactgttcttctagtgtagccgtagttaggccaccacttcaagaactctgtagcaccgcctacatacctcgctctgctaatcctgttaccagtggctgctgccagtggcgataagtcgtgtcttaccgggttggactcaagacgatagttaccggataaggcgcagcggtcgggctgaacggggggttcgtgcacacagcccagcttggagcgaacgacctacaccgaactgagatacctacagcgtgagctatgagaaagcgccacgcttcccgaagggagaaaggcggacaggtatccggtaagcggcagggtcggaacaggagagcgcacgagggagcttccagggggaaacgcctggtatctttatagtcctgtcgggtttcgccacctctgacttgagcgtcgatttttgtgatgctcgtcaggggggcggagcctatggaaaaacgccagcaacgcggcctttttacggttcctggccttttgctggccttttgctcacatgttctttcctgcgttatcccctgattctgtggataaccgtattaccgctagcatggatctcggggacgtctaactactaagcgagagtagggaactgccaggcatcaaataaaacgaaaggctcagtcggaagactgggcctttcgttttatctgttgtttgtcggtgaacgctctcctgagtaggacaaatccgccgggagcggatttgaacgttgtgaagcaacggcccggagggtggcgggcaggacgcccgccataaactgccaggcatcaaactaagcagaaggccatcctgacggatggcctttttgcgtttctacaaactcttcctgttagttagttacttaagcgccagggttttcccagtcacgacgttgtaaaacgacggccagtgcatacgatttaggtgacactatagaagggccccaaataatgattttattttgactgatagtgacctgttcgttgcaacaaattgataagcaatgcttttttataatgccaactttgtacaaaaaagcaggctttactgaagcttcggaattcggcgcgccagtgaattatcaactatgtataataaagttgaacgagaaacgtaaaatgatataaatatcaatatattaaattagattttgcataaaaaacagactacataatactgtaaaacacaacatatccagtcactatggcggccgttcggggaaatgtgcgcggaacccctatttgtttatttttctaaatacattcaaatatgtatccgctcatgagacaataaccctgataaatgcttcaataatattgaaaaaggaagagtatgagtattcaacatttccgtgtcgcccttattcccttttttgcggcattttgccttcctgtttttgctcacccagaaacgctggtgaaagtaaaagatgctgaagatcagttgggtgcacgagtgggttacatcgaactggatctcaacagcggtaagatccttgagagttttcgccccgaagaacgttttccaatgatgagcacttttaaagttctgctatgtggcgcggtattatcccgtattgacgccgggcaagagcaactcggtcgccgcatacactattctcagaatgacttggttgagtactcaccagtcacagaaaagcatcttacggatggcatgacagtaagagaattatgcagtgctgccataaccatgagtgataacactgcggccaacttacttctgacaacgatcggaggaccgaaggagcta
Vector information for pL34R12-Cm
Map of pL34R12-Cm
Location of Features
•Cm(R):3408-406
•attR2: 548-672 (c)
•attL4: 692-787 (c)
•T7:807-826
•RV-M:851-874
•Gen(R):1136-1669
•pUCorigin: 1762-2435
•rrnB T1: 2544-2587 (c)
•rrnB T2: 2719-2762(c)
•M13-47:2787-2810 (c)
•SP6:2832-2855 (c)
•attL3: 2876-2971
•attR1: 3004-3128
(c) = complementary strand
Sequence of pL34R12-Cm
ggatagtgttcacccttgttacaccgttttccatgagcaaactgaaacgttttcatcgctctggagtgaataccacgacgatttccggcagtttctacacatatattcgcaagatgtggcgtgttacggtgaaaacctggcctatttccctaaagggtttattgagaatatgtttttcgtctcagccaatccctgggtgagtttcaccagttttgatttaaacgtggccaatatggacaacttcttcgcccccgttttcacaatgggcaaatattatacgcaaggcgacaaggtgctgatgccgctggcgattcaggttcatcatgccgtttgtgatggcttccatgtcggcagaatgcttaatgaattacaacagtactgcgatgagtggcagggcggggcgtaatttttttaaggcagttattggtgcgatcctctagctagaaattcgttcaagccgacgccgcttcgccggcgttaactcaagcgattagatgcactaagcacataattgctcacagccaaactatcaggtcaagtcctcgaccatagtgactggatatgttgtgttttacagtattatgtagtctgttttttatgcaaaatctaatttaatatattgatatttatatcattttacgtttctcgttcagctttcttgtacaaagtggttcgataattccttaattaacaacttttctatacaaagttggcattataaaaaagcattgcttatcaatttgttgcaacgaacaggtcactatcagtcaaaataaaatcattatttggaatcgtcgagctgcaggccctatagtgagtcgtattaccgtaatcatggtcatagctgtttcctgtgtgaaattgttatccgctctcatgagacgcacaccgtggaaacggatgaaggcacgaacccagttgacataagcctgttcggttcgtaaactgtaatgcaagtagcgtatgcgctcacgcaactggtccagaaccttgaccgaacgcagcggtggtaacggcgcagtggcggttttcatggcttgttatgactgtttttttgtacagtctatgcctcgggcatccaagcagcaagcgcgttacgccgtgggtcgatgtttgatgttatggagcagcaacgatgttacgcagcagcaacgatgttacgcagcagggcagtcgccctaaaacaaagttaggtggctcaagtatgggcatcattcgcacatgtaggctcggccctgaccaagtcaaatccatgcgggctgctcttgatcttttcggtcgtgagttcggagacgtagccacctactcccaacatcagccggactccgattacctcgggaacttgctccgtagtaagacattcatcgcgcttgctgccttcgaccaagaagcggttgttggcgctctcgcggcttacgttctgcccaggtttgagcagccgcgtagtgagatctatatctatgatctcgcagtctccggcgagcaccggaggcagggcattgccaccgcgctcatcaatctcctcaagcatgaggccaacgcgcttggtgcttatgtgatctacgtgcaagcagattacggtgacgatcccgcagtggctctctatacaaagttgggcatacgggaagaagtgatgcactttgatatcgacccaagtaccgccacctaacaattcgttcaagccgagatcggcttcccggccgcggagttgttcggtaaattgtcacaacgccgcccgccccgttccactgagcgtcagacccggtagaaaagatcaaaggatcttcttgagatcctttttttctgcgcgtaatctgctgcttgcaaacaaaaaaaccaccgctaccagcggtggtttgtttgccggatcaagagctaccaactctttttccgaaggtaactggcttcagcagagcgcagataccaaatactgttcttctagtgtagccgtagttaggccaccacttcaagaactctgtagcaccgcctacatacctcgctctgctaatcctgttaccagtggctgctgccagtggcgataagtcgtgtcttaccgggttggactcaagacgatagttaccggataaggcgcagcggtcgggctgaacggggggttcgtgcacacagcccagcttggagcgaacgacctacaccgaactgagatacctacagcgtgagctatgagaaagcgccacgcttcccgaagggagaaaggcggacaggtatccggtaagcggcagggtcggaacaggagagcgcacgagggagcttccagggggaaacgcctggtatctttatagtcctgtcgggtttcgccacctctgacttgagcgtcgatttttgtgatgctcgtcaggggggcggagcctatggaaaaacgccagcaacgcggcctttttacggttcctggccttttgctggccttttgctcacatgttctttcctgcgttatcccctgattctgtggataaccgtattaccgctagcatggatctcggggacgtctaactactaagcgagagtagggaactgccaggcatcaaataaaacgaaaggctcagtcggaagactgggcctttcgttttatctgttgtttgtcggtgaacgctctcctgagtaggacaaatccgccgggagcggatttgaacgttgtgaagcaacggcccggagggtggcgggcaggacgcccgccataaactgccaggcatcaaactaagcagaaggccatcctgacggatggcctttttgcgtttctacaaactcttcctgttagttagttacttaagcgccagggttttcccagtcacgacgttgtaaaacgacggccagtgcatacgatttaggtgacactatagaagggcccactagagattccaaataatgattttattttgactgatagtgacctgttcgttgcaacaaattgataagcaatgcttttttataatgccaactttgtataataaagttgaagcttcggaattcggcgcgccaagctatcaaacaagtttgtacaaaaaagctgaacgagaaacgtaaaatgatataaatatcaatatattaaattagattttgcataaaaaacagactacataatactgtaaaacacaacatatccagtcactatggcggccggtggacataagcctcgttcggttcgtaagctgtaatgcaagtagcgtaactgccgtcacgcaactggtccagaaccttgaccgaacgcagcggtggtaacggcgcagtggcggttttcatggcttcttgttatgacatgtttttttggggtacagtctatgcctcgggcatccaagcagcaagcgcgttacgccgtgggtcgatgtttgatgttatggagcagcaacgatgttacgcagcagggcagtcgccctaaaacaaagttaaacatcatggagaaaaaaatcactggatataccaccgttgatatatcccaatggcatcgtaaagaacattttgaggcatttcagtcagttgctcaatgtacctataaccagaccgttcagctggatattacggcctttttaaagaccgtaaagaaaaataagcacaagttttatccggcctttattcacattcttgcccgcctgatgaatgctcatccggagttccgtatggcaatgaaagacggtgagctggtgatatg
Vector information for pL12R34-sacB
Map of pL12R34-sacB
Location of Features
•sacB: 4008-915
•attR4: 964-1088 (c)
•attL2: 1121-1220 (c)
•T7: 1236-1255 (c)
•RV-M: 1280-1303 (c)
•Gen(R): 1565-2098
•pUCorigin: 2191-2864
•rrnB T1: 2973-3016 (c)
•rrnB T2: 3148-3191 (c)
•M13-47: 3216-3239
•SP6: 3261-3284
•attL1: 3293-3392
•attR3: 3431-3555
(c) = complementary strand
Sequence of pL12R34-sacB
tttacatctgacggaaaaatccgtttattctacactgatttctccggtaaacattacggcaaacaaacactgacaactgcacaagttaacgtatcagcatcagacagctctttgaacatcaacggtgtagaggattataaatcaatctttgacggtgacggaaaaacgtatcaaaatgtacagcagttcatcgatgaaggcaactacagctcaggcgacaaccatacgctgagagatcctcactacgtagaagataaaggccacaaatacttagtatttgaagcaaacactggaactgaagatggctaccaaggcgaagaatctttatttaacaaagcatactatggcaaaagcacatcattcttccgtcaagaaagtcaaaaacttctgcaaagcgataaaaaacgcacggctgagttagcaaacggcgctctcggtatgattgagctaaacgatgattacacactgaaaaaagtgatgaaaccgctgattgcatctaacacagtaacagatgaaattgaacgcgcgaacgtctttaaaatgaacggcaaatggtacctgttcactgactcccgcggatcaaaaatgacgattgacggcattacgtctaacgatatttacatgcttggttatgtttctaattctttaactggcccatacaagccgctgaacaaaactggccttgtgttaaaaatggatcttgatcctaacgatgtaacctttacttactcacacttcgctgtacctcaagcgaaaggaaacaatgtcgtgattacaagctatatgacaaacagaggattctacgcagacaaacaatcaacgtttgcgcctagcttcctgctgaacatcaaaggcaagaaaacatctgttgtcaaagacagcatccttgaacaaggacaattaacagttaacaaataaaaacgcaaaagaaaatgccgatatcctattggcattgtcgatcgaccatagtgactggatatgttgtgttttacagtattatgtagtctgttttttatgcaaaatctaatttaatatattgatatttatatcattttacgtttctcgttcaacttttctatacaaagttgatagcttggcgtaatcatgttaattaagctagacccagctttcttgtacaaagttggcattataagaaagcattgcttatcaatttgttgcaacgaacaggtcactatcagtcaaaataaaatcattatttgccatccagctgcaggccctatagtgagtcgtattaccgtaatcatggtcatagctgtttcctgtgtgaaattgttatccgctctcatgagacgcacaccgtggaaacggatgaaggcacgaacccagttgacataagcctgttcggttcgtaaactgtaatgcaagtagcgtatgcgctcacgcaactggtccagaaccttgaccgaacgcagcggtggtaacggcgcagtggcggttttcatggcttgttatgactgtttttttgtacagtctatgcctcgggcatccaagcagcaagcgcgttacgccgtgggtcgatgtttgatgttatggagcagcaacgatgttacgcagcagcaacgatgttacgcagcagggcagtcgccctaaaacaaagttaggtggctcaagtatgggcatcattcgcacatgtaggctcggccctgaccaagtcaaatccatgcgggctgctcttgatcttttcggtcgtgagttcggagacgtagccacctactcccaacatcagccggactccgattacctcgggaacttgctccgtagtaagacattcatcgcgcttgctgccttcgaccaagaagcggttgttggcgctctcgcggcttacgttctgcccaggtttgagcagccgcgtagtgagatctatatctatgatctcgcagtctccggcgagcaccggaggcagggcattgccaccgcgctcatcaatctcctcaagcatgaggccaacgcgcttggtgcttatgtgatctacgtgcaagcagattacggtgacgatcccgcagtggctctctatacaaagttgggcatacgggaagaagtgatgcactttgatatcgacccaagtaccgccacctaacaattcgttcaagccgagatcggcttcccggccgcggagttgttcggtaaattgtcacaacgccgcccgccccgttccactgagcgtcagacccggtagaaaagatcaaaggatcttcttgagatcctttttttctgcgcgtaatctgctgcttgcaaacaaaaaaaccaccgctaccagcggtggtttgtttgccggatcaagagctaccaactctttttccgaaggtaactggcttcagcagagcgcagataccaaatactgttcttctagtgtagccgtagttaggccaccacttcaagaactctgtagcaccgcctacatacctcgctctgctaatcctgttaccagtggctgctgccagtggcgataagtcgtgtcttaccgggttggactcaagacgatagttaccggataaggcgcagcggtcgggctgaacggggggttcgtgcacacagcccagcttggagcgaacgacctacaccgaactgagatacctacagcgtgagctatgagaaagcgccacgcttcccgaagggagaaaggcggacaggtatccggtaagcggcagggtcggaacaggagagcgcacgagggagcttccagggggaaacgcctggtatctttatagtcctgtcgggtttcgccacctctgacttgagcgtcgatttttgtgatgctcgtcaggggggcggagcctatggaaaaacgccagcaacgcggcctttttacggttcctggccttttgctggccttttgctcacatgttctttcctgcgttatcccctgattctgtggataaccgtattaccgctagcatggatctcggggacgtctaactactaagcgagagtagggaactgccaggcatcaaataaaacgaaaggctcagtcggaagactgggcctttcgttttatctgttgtttgtcggtgaacgctctcctgagtaggacaaatccgccgggagcggatttgaacgttgtgaagcaacggcccggagggtggcgggcaggacgcccgccataaactgccaggcatcaaactaagcagaaggccatcctgacggatggcctttttgcgtttctacaaactcttcctgttagttagttacttaagcgccagggttttcccagtcacgacgttgtaaaacgacggccagtgcatacgatttaggtgacactatagaagggccccaaataatgattttattttgactgatagtgacctgttcgttgcaacaaattgataagcaatgcttttttataatgccaactttgtacaaaaaagcaggctttactgaagcttcggaattcggcgcgccagtgaattatcaactatgtataataaagttgaacgagaaacgtaaaatgatataaatatcaatatattaaattagattttgcataaaaaacagactacataatactgtaaaacacaacatatccagtcactatggcggccccacatatacctgccgttcactattatttagtgaaatgagatattatgatattttctgaattgtgattaaaaaggcaactttatgcccatgcaacagaaactataaaaaatacagagaatgaaaagaaacagatagattttttagttctttaggcccgtagtctgcaaatccttttatgattttctatcaaacaaaagaggaaaatagaccagttgcaatccaaacgagagtctaatagaatgaggtcgaaaagtaaatcgcgcgggtttgttactgataaagcaggcaagacctaaaatgtgtaaagggcaaagtgtatactttggcgtcaccccttacatattttaggtctttttttattgtgcgtaactaacttgccatcttcaaacaggagggctggaagaagcagaccgctaacacagtacataaaaaaggagacatgaacgatgaacatcaaaaagtttgcaaaacaagcaacagtattaacctttactaccgcactgctggcaggaggcgcaactcaagcgtttgcgaaagaaacgaaccaaaagccatataaggaaacatacggcatttcccatattacacgccatgatatgctgcaaatccctgaacagcaaaaaaatgaaaaatatcaagttcctgagttcgattcgtccacaattaaaaatatctcttctgcaaaaggcctggacgtttgggacagctggccattacaaaacgctgacggcactgtcgcaaactatcacggctaccacatcgtctttgcattagccggagatcctaaaaatgcggatgacacatcgatttacatgttctatcaaaaagtcggcgaaacttctattgacagctggaaaaacgctggccgcgtctttaaagacagcgacaaattcgatgcaaatgattctatcctaaaagaccaaacacaagaatggtcaggttcagccaca
Vector information for pL34R12-ccdB
Map of pL34R12-ccdB
Location of Features
•ccdB: 5-310
•attR2: 351-475 (c)
•attL4: 495-590 (c)
•T7: 610-629
•RV-M: 654-677
•Gen(R): 939-1472
•pUCorigin: 1565-2238
•rrnB T1: 2347-2390 (c)
•rrnB T2: 2522-2565(c)
•M13-47: 2590-2613 (c)
•SP6: 2635-2658 (c)
•attL3: 2679-2774
•attR1: 2807-2931
(c) = complementary strand
Sequence of pL34R12-ccdB
tgaaatgcagtttaaggtttacacctataaaagagagagccgttatcgtctgtttgtggatgtacagagtgatattattgacacgcccgggcgacggatggtgatccccctggccagtgcacgtctgctgtcagataaagtctcccgtgaactttacccggtggtgcatatcggggatgaaagctggcgcatgatgaccaccgatatggccagtgtgccggtctccgttatcggggaagaagtggctgatctcagccaccgcgaaaatgacatcaaaaacgccattaacctgatgttctggggaatataaatgtcaggctcccttatacacagccagtctgcaggtcgaccatagtgactggatatgttgtgttttacagtattatgtagtctgttttttatgcaaaatctaatttaatatattgatatttatatcattttacgtttctcgttcagctttcttgtacaaagtggttcgataattccttaattaacaacttttctatacaaagttggcattataaaaaagcattgcttatcaatttgttgcaacgaacaggtcactatcagtcaaaataaaatcattatttggaatcgtcgagctgcaggccctatagtgagtcgtattaccgtaatcatggtcatagctgtttcctgtgtgaaattgttatccgctctcatgagacgcacaccgtggaaacggatgaaggcacgaacccagttgacataagcctgttcggttcgtaaactgtaatgcaagtagcgtatgcgctcacgcaactggtccagaaccttgaccgaacgcagcggtggtaacggcgcagtggcggttttcatggcttgttatgactgtttttttgtacagtctatgcctcgggcatccaagcagcaagcgcgttacgccgtgggtcgatgtttgatgttatggagcagcaacgatgttacgcagcagcaacgatgttacgcagcagggcagtcgccctaaaacaaagttaggtggctcaagtatgggcatcattcgcacatgtaggctcggccctgaccaagtcaaatccatgcgggctgctcttgatcttttcggtcgtgagttcggagacgtagccacctactcccaacatcagccggactccgattacctcgggaacttgctccgtagtaagacattcatcgcgcttgctgccttcgaccaagaagcggttgttggcgctctcgcggcttacgttctgcccaggtttgagcagccgcgtagtgagatctatatctatgatctcgcagtctccggcgagcaccggaggcagggcattgccaccgcgctcatcaatctcctcaagcatgaggccaacgcgcttggtgcttatgtgatctacgtgcaagcagattacggtgacgatcccgcagtggctctctatacaaagttgggcatacgggaagaagtgatgcactttgatatcgacccaagtaccgccacctaacaattcgttcaagccgagatcggcttcccggccgcggagttgttcggtaaattgtcacaacgccgcccgccccgttccactgagcgtcagacccggtagaaaagatcaaaggatcttcttgagatcctttttttctgcgcgtaatctgctgcttgcaaacaaaaaaaccaccgctaccagcggtggtttgtttgccggatcaagagctaccaactctttttccgaaggtaactggcttcagcagagcgcagataccaaatactgttcttctagtgtagccgtagttaggccaccacttcaagaactctgtagcaccgcctacatacctcgctctgctaatcctgttaccagtggctgctgccagtggcgataagtcgtgtcttaccgggttggactcaagacgatagttaccggataaggcgcagcggtcgggctgaacggggggttcgtgcacacagcccagcttggagcgaacgacctacaccgaactgagatacctacagcgtgagctatgagaaagcgccacgcttcccgaagggagaaaggcggacaggtatccggtaagcggcagggtcggaacaggagagcgcacgagggagcttccagggggaaacgcctggtatctttatagtcctgtcgggtttcgccacctctgacttgagcgtcgatttttgtgatgctcgtcaggggggcggagcctatggaaaaacgccagcaacgcggcctttttacggttcctggccttttgctggccttttgctcacatgttctttcctgcgttatcccctgattctgtggataaccgtattaccgctagcatggatctcggggacgtctaactactaagcgagagtagggaactgccaggcatcaaataaaacgaaaggctcagtcggaagactgggcctttcgttttatctgttgtttgtcggtgaacgctctcctgagtaggacaaatccgccgggagcggatttgaacgttgtgaagcaacggcccggagggtggcgggcaggacgcccgccataaactgccaggcatcaaactaagcagaaggccatcctgacggatggcctttttgcgtttctacaaactcttcctgttagttagttacttaagcgccagggttttcccagtcacgacgttgtaaaacgacggccagtgcatacgatttaggtgacactatagaagggcccactagagattccaaataatgattttattttgactgatagtgacctgttcgttgcaacaaattgataagcaatgcttttttataatgccaactttgtataataaagttgaagcttcggaattcggcgcgccaagctatcaaacaagtttgtacaaaaaagctgaacgagaaacgtaaaatgatataaatatcaatatattaaattagattttgcataaaaaacagactacataatactgtaaaacacaacatatccagtcactatggcggccgatccggcttactaaaagccagataacagtatgcgtatttgcgcgctgatttttgcggtataagaatatatactgatatgtatacccgaagtatgtcaaaaagaggtgtgctatgaagcagcgtattacagtgacagttgacagcgacagctatcagttgctcaaggcatatatgatgtcaatatctccggtctggtaagcacaaccatgcagaatgaagcccgtcgtctgcgtgccgaacgctggaaagcggaaaatcaggaagggatggctgaggtcgcccggtttattgaaatgaacggctcttttgctgacgagaacagggactgg
Vector information for pL12R34H-Ap
Map of pL12R34H-Ap
Location of Features
•Ap(R): 3259-423
•PI-SceI(R): 565-603 (c)
•attR4: 608-732 (c)
•attL2: 765-864 (c)
•I-SceI(R): 879-896
•Gen(R): 1158-1691
•pUCorigin: 1784-2457
•rrnB T1: 2566-2609 (c)
•rrnB T2: 2741-2784(c)
•I-SceI: 2809-2826 (c)
•attL1: 2833-2932
•attR3: 2971-3095
•PI-SceI: 3101-3139
(c) = complementary strand
Sequence of pL12R34H-Ap
accgcttttttgcacaacatgggggatcatgtaactcgccttgatcgttgggaaccggagctgaatgaagccataccaaacgacgagcgtgacaccacgatgcctgtagcaatggcaacaacgttgcgcaaactattaactggcgaactacttactctagcttcccggcaacaattaatagactggatggaggcggataaagttgcaggaccacttctgcgctcggcccttccggctggctggtttattgctgataaatctggagccggtgagcgtgggtctcgcggtatcattgcagcactggggccagatggtaagccctcccgtatcgtagttatctacacgacggggagtcaggcaactatggatgaacgaaatagacagatcgctgagataggtgcctcactgattaagcattggtaactgtcagaccaagtttactcatatatactttagattgatttaaaacttcatttttaatttaaaaggatctaggtgaagatcctttttgataatctcatgaccaaaatcccttaacgtgagttttcgttccactgagcgtcatgccatttcattacctctttctccgcacccgacatagattcgaccatagtgactggatatgttgtgttttacagtattatgtagtctgttttttatgcaaaatctaatttaatatattgatatttatatcattttacgtttctcgttcaacttttctatacaaagttgatagcttggcgtaatcatgttaattaagctagacccagctttcttgtacaaagttggcattataagaaagcattgcttatcaatttgttgcaacgaacaggtcactatcagtcaaaataaaatcattatttgccatccagctgcacattaccctgttatccctaccatgagacgcacaccgtggaaacggatgaaggcacgaacccagttgacataagcctgttcggttcgtaaactgtaatgcaagtagcgtatgcgctcacgcaactggtccagaaccttgaccgaacgcagcggtggtaacggcgcagtggcggttttcatggcttgttatgactgtttttttgtacagtctatgcctcgggcatccaagcagcaagcgcgttacgccgtgggtcgatgtttgatgttatggagcagcaacgatgttacgcagcagcaacgatgttacgcagcagggcagtcgccctaaaacaaagttaggtggctcaagtatgggcatcattcgcacatgtaggctcggccctgaccaagtcaaatccatgcgggctgctcttgatcttttcggtcgtgagttcggagacgtagccacctactcccaacatcagccggactccgattacctcgggaacttgctccgtagtaagacattcatcgcgcttgctgccttcgaccaagaagcggttgttggcgctctcgcggcttacgttctgcccaggtttgagcagccgcgtagtgagatctatatctatgatctcgcagtctccggcgagcaccggaggcagggcattgccaccgcgctcatcaatctcctcaagcatgaggccaacgcgcttggtgcttatgtgatctacgtgcaagcagattacggtgacgatcccgcagtggctctctatacaaagttgggcatacgggaagaagtgatgcactttgatatcgacccaagtaccgccacctaacaattcgttcaagccgagatcggcttcccggccgcggagttgttcggtaaattgtcacaacgccgcccgccccgttccactgagcgtcagacccggtagaaaagatcaaaggatcttcttgagatcctttttttctgcgcgtaatctgctgcttgcaaacaaaaaaaccaccgctaccagcggtggtttgtttgccggatcaagagctaccaactctttttccgaaggtaactggcttcagcagagcgcagataccaaatactgttcttctagtgtagccgtagttaggccaccacttcaagaactctgtagcaccgcctacatacctcgctctgctaatcctgttaccagtggctgctgccagtggcgataagtcgtgtcttaccgggttggactcaagacgatagttaccggataaggcgcagcggtcgggctgaacggggggttcgtgcacacagcccagcttggagcgaacgacctacaccgaactgagatacctacagcgtgagctatgagaaagcgccacgcttcccgaagggagaaaggcggacaggtatccggtaagcggcagggtcggaacaggagagcgcacgagggagcttccagggggaaacgcctggtatctttatagtcctgtcgggtttcgccacctctgacttgagcgtcgatttttgtgatgctcgtcaggggggcggagcctatggaaaaacgccagcaacgcggcctttttacggttcctggccttttgctggccttttgctcacatgttctttcctgcgttatcccctgattctgtggataaccgtattaccgctagcatggatctcggggacgtctaactactaagcgagagtagggaactgccaggcatcaaataaaacgaaaggctcagtcggaagactgggcctttcgttttatctgttgtttgtcggtgaacgctctcctgagtaggacaaatccgccgggagcggatttgaacgttgtgaagcaacggcccggagggtggcgggcaggacgcccgccataaactgccaggcatcaaactaagcagaaggccatcctgacggatggcctttttgcgtttctacaaactcttcctgttagttagttacttaactagggataacagggtaatcggccccaaataatgattttattttgactgatagtgacctgttcgttgcaacaaattgataagcaatgcttttttataatgccaactttgtacaaaaaagcaggctttactgaagcttcggaattcggcgcgccagtgaattatcaactatgtataataaagttgaacgagaaacgtaaaatgatataaatatcaatatattaaattagattttgcataaaaaacagactacataatactgtaaaacacaacatatccagtcactatggcggccatctatgtcgggtgcggagaaagaggtaatgaaatggcattcggggaaatgtgcgcggaacccctatttgtttatttttctaaatacattcaaatatgtatccgctcatgagacaataaccctgataaatgcttcaataatattgaaaaaggaagagtatgagtattcaacatttccgtgtcgcccttattcccttttttgcggcattttgccttcctgtttttgctcacccagaaacgctggtgaaagtaaaagatgctgaagatcagttgggtgcacgagtgggttacatcgaactggatctcaacagcggtaagatccttgagagttttcgccccgaagaacgttttccaatgatgagcacttttaaagttctgctatgtggcgcggtattatcccgtattgacgccgggcaagagcaactcggtcgccgcatacactattctcagaatgacttggttgagtactcaccagtcacagaaaagcatcttacggatggcatgacagtaagagaattatgcagtgctgccataaccatgagtgataacactgcggccaacttacttctgacaacgatcggaggaccgaaggagcta
Vector information for pL34R12H-Cm
Map of pL34R12H-Cm
Location of Features
•Cm(R): 3371-406
•PI-SceI(R): 537-575(c)
•attR2: 581-705(c)
•attL4: 725-820(c)
•I-SceI(R): 839-856(c)
•Gen(R): 1118-1651
•pUCorigin: 1744-2417
•rrnB T1: 2526-2569 (c)
•rrnB T2: 2701-2744 (c)
•I-SceI: 2769-2786
•attL3: 2805-2900
•attR1: 2933-3057
•PI-SceI: 3064-3102
(c) = complementary strand
Sequence of pL34R12H-Cm
ggatagtgttcacccttgttacaccgttttccatgagcaaactgaaacgttttcatcgctctggagtgaataccacgacgatttccggcagtttctacacatatattcgcaagatgtggcgtgttacggtgaaaacctggcctatttccctaaagggtttattgagaatatgtttttcgtctcagccaatccctgggtgagtttcaccagttttgatttaaacgtggccaatatggacaacttcttcgcccccgttttcacaatgggcaaatattatacgcaaggcgacaaggtgctgatgccgctggcgattcaggttcatcatgccgtttgtgatggcttccatgtcggcagaatgcttaatgaattacaacagtactgcgatgagtggcagggcggggcgtaatttttttaaggcagttattggtgcgatcctctagctagaaattcgttcaagccgacgccgcttcgccggcgttaactcaagcgattagatgcactaagcacataattgctcacagccaaactatcaggtctgccatttcattacctctttctccgcacccgacatagattcgaccatagtgactggatatgttgtgttttacagtattatgtagtctgttttttatgcaaaatctaatttaatatattgatatttatatcattttacgtttctcgttcagctttcttgtacaaagtggttcgataattccttaattaacaacttttctatacaaagttggcattataaaaaagcattgcttatcaatttgttgcaacgaacaggtcactatcagtcaaaataaaatcattatttggaatcgtcgagctgcacattaccctgttatccctaccatgagacgcacaccgtggaaacggatgaaggcacgaacccagttgacataagcctgttcggttcgtaaactgtaatgcaagtagcgtatgcgctcacgcaactggtccagaaccttgaccgaacgcagcggtggtaacggcgcagtggcggttttcatggcttgttatgactgtttttttgtacagtctatgcctcgggcatccaagcagcaagcgcgttacgccgtgggtcgatgtttgatgttatggagcagcaacgatgttacgcagcagcaacgatgttacgcagcagggcagtcgccctaaaacaaagttaggtggctcaagtatgggcatcattcgcacatgtaggctcggccctgaccaagtcaaatccatgcgggctgctcttgatcttttcggtcgtgagttcggagacgtagccacctactcccaacatcagccggactccgattacctcgggaacttgctccgtagtaagacattcatcgcgcttgctgccttcgaccaagaagcggttgttggcgctctcgcggcttacgttctgcccaggtttgagcagccgcgtagtgagatctatatctatgatctcgcagtctccggcgagcaccggaggcagggcattgccaccgcgctcatcaatctcctcaagcatgaggccaacgcgcttggtgcttatgtgatctacgtgcaagcagattacggtgacgatcccgcagtggctctctatacaaagttgggcatacgggaagaagtgatgcactttgatatcgacccaagtaccgccacctaacaattcgttcaagccgagatcggcttcccggccgcggagttgttcggtaaattgtcacaacgccgcccgccccgttccactgagcgtcagacccggtagaaaagatcaaaggatcttcttgagatcctttttttctgcgcgtaatctgctgcttgcaaacaaaaaaaccaccgctaccagcggtggtttgtttgccggatcaagagctaccaactctttttccgaaggtaactggcttcagcagagcgcagataccaaatactgttcttctagtgtagccgtagttaggccaccacttcaagaactctgtagcaccgcctacatacctcgctctgctaatcctgttaccagtggctgctgccagtggcgataagtcgtgtcttaccgggttggactcaagacgatagttaccggataaggcgcagcggtcgggctgaacggggggttcgtgcacacagcccagcttggagcgaacgacctacaccgaactgagatacctacagcgtgagctatgagaaagcgccacgcttcccgaagggagaaaggcggacaggtatccggtaagcggcagggtcggaacaggagagcgcacgagggagcttccagggggaaacgcctggtatctttatagtcctgtcgggtttcgccacctctgacttgagcgtcgatttttgtgatgctcgtcaggggggcggagcctatggaaaaacgccagcaacgcggcctttttacggttcctggccttttgctggccttttgctcacatgttctttcctgcgttatcccctgattctgtggataaccgtattaccgctagcatggatctcggggacgtctaactactaagcgagagtagggaactgccaggcatcaaataaaacgaaaggctcagtcggaagactgggcctttcgttttatctgttgtttgtcggtgaacgctctcctgagtaggacaaatccgccgggagcggatttgaacgttgtgaagcaacggcccggagggtggcgggcaggacgcccgccataaactgccaggcatcaaactaagcagaaggccatcctgacggatggcctttttgcgtttctacaaactcttcctgttagttagttacttaactagggataacagggtaatcggcccactagagattccaaataatgattttattttgactgatagtgacctgttcgttgcaacaaattgataagcaatgcttttttataatgccaactttgtataataaagttgaagcttcggaattcggcgcgccaagctatcaaacaagtttgtacaaaaaagctgaacgagaaacgtaaaatgatataaatatcaatatattaaattagattttgcataaaaaacagactacataatactgtaaaacacaacatatccagtcactatggcggccatctatgtcgggtgcggagaaagaggtaatgaaatggcaacataagcctcgttcggttcgtaagctgtaatgcaagtagcgtaactgccgtcacgcaactggtccagaaccttgaccgaacgcagcggtggtaacggcgcagtggcggttttcatggcttcttgttatgacatgtttttttggggtacagtctatgcctcgggcatccaagcagcaagcgcgttacgccgtgggtcgatgtttgatgttatggagcagcaacgatgttacgcagcagggcagtcgccctaaaacaaagttaaacatcatggagaaaaaaatcactggatataccaccgttgatatatcccaatggcatcgtaaagaacattttgaggcatttcagtcagttgctcaatgtacctataaccagaccgttcagctggatattacggcctttttaaagaccgtaaagaaaaataagcacaagttttatccggcctttattcacattcttgcccgcctgatgaatgctcatccggagttccgtatggcaatgaaagacggtgagctggtgatatg
Vector information for pL12R34H-sacB
Map of pL12R34H-sacB
Location of Features
•sacB: 3971-942
•attR4: 985-1109(c)
•attL2: 1142-1241(c)
•I-SceI(R): 1256-1273(c)
•Gen(R): 1535-2068
•pUCorigin: 2161-2834
•rrnB T1: 2943-2986 (c)
•rrnB T2: 3118-3161 (c)
•I-SceI: 3186-3203
•attL1: 3210-3309
•attR3: 3348-3472
•PI-SceI(R): 3480-3518(c)
(c) = complementary strand
Sequence of pL12R34H-sacB
caagaatggtcaggttcagccacatttacatctgacggaaaaatccgtttattctacactgatttctccggtaaacattacggcaaacaaacactgacaactgcacaagttaacgtatcagcatcagacagctctttgaacatcaacggtgtagaggattataaatcaatctttgacggtgacggaaaaacgtatcaaaatgtacagcagttcatcgatgaaggcaactacagctcaggcgacaaccatacgctgagagatcctcactacgtagaagataaaggccacaaatacttagtatttgaagcaaacactggaactgaagatggctaccaaggcgaagaatctttatttaacaaagcatactatggcaaaagcacatcattcttccgtcaagaaagtcaaaaacttctgcaaagcgataaaaaacgcacggctgagttagcaaacggcgctctcggtatgattgagctaaacgatgattacacactgaaaaaagtgatgaaaccgctgattgcatctaacacagtaacagatgaaattgaacgcgcgaacgtctttaaaatgaacggcaaatggtacctgttcactgactcccgcggatcaaaaatgacgattgacggcattacgtctaacgatatttacatgcttggttatgtttctaattctttaactggcccatacaagccgctgaacaaaactggccttgtgttaaaaatggatcttgatcctaacgatgtaacctttacttactcacacttcgctgtacctcaagcgaaaggaaacaatgtcgtgattacaagctatatgacaaacagaggattctacgcagacaaacaatcaacgtttgcgcctagcttcctgctgaacatcaaaggcaagaaaacatctgttgtcaaagacagcatccttgaacaaggacaattaacagttaacaaataaaaacgcaaaagaaaatgccgatatcctattggcattgctcgaccatagtgactggatatgttgtgttttacagtattatgtagtctgttttttatgcaaaatctaatttaatatattgatatttatatcattttacgtttctcgttcaacttttctatacaaagttgatagcttggcgtaatcatgttaattaagctagacccagctttcttgtacaaagttggcattataagaaagcattgcttatcaatttgttgcaacgaacaggtcactatcagtcaaaataaaatcattatttgccatccagctgcacattaccctgttatccctaccatgagacgcacaccgtggaaacggatgaaggcacgaacccagttgacataagcctgttcggttcgtaaactgtaatgcaagtagcgtatgcgctcacgcaactggtccagaaccttgaccgaacgcagcggtggtaacggcgcagtggcggttttcatggcttgttatgactgtttttttgtacagtctatgcctcgggcatccaagcagcaagcgcgttacgccgtgggtcgatgtttgatgttatggagcagcaacgatgttacgcagcagcaacgatgttacgcagcagggcagtcgccctaaaacaaagttaggtggctcaagtatgggcatcattcgcacatgtaggctcggccctgaccaagtcaaatccatgcgggctgctcttgatcttttcggtcgtgagttcggagacgtagccacctactcccaacatcagccggactccgattacctcgggaacttgctccgtagtaagacattcatcgcgcttgctgccttcgaccaagaagcggttgttggcgctctcgcggcttacgttctgcccaggtttgagcagccgcgtagtgagatctatatctatgatctcgcagtctccggcgagcaccggaggcagggcattgccaccgcgctcatcaatctcctcaagcatgaggccaacgcgcttggtgcttatgtgatctacgtgcaagcagattacggtgacgatcccgcagtggctctctatacaaagttgggcatacgggaagaagtgatgcactttgatatcgacccaagtaccgccacctaacaattcgttcaagccgagatcggcttcccggccgcggagttgttcggtaaattgtcacaacgccgcccgccccgttccactgagcgtcagacccggtagaaaagatcaaaggatcttcttgagatcctttttttctgcgcgtaatctgctgcttgcaaacaaaaaaaccaccgctaccagcggtggtttgtttgccggatcaagagctaccaactctttttccgaaggtaactggcttcagcagagcgcagataccaaatactgttcttctagtgtagccgtagttaggccaccacttcaagaactctgtagcaccgcctacatacctcgctctgctaatcctgttaccagtggctgctgccagtggcgataagtcgtgtcttaccgggttggactcaagacgatagttaccggataaggcgcagcggtcgggctgaacggggggttcgtgcacacagcccagcttggagcgaacgacctacaccgaactgagatacctacagcgtgagctatgagaaagcgccacgcttcccgaagggagaaaggcggacaggtatccggtaagcggcagggtcggaacaggagagcgcacgagggagcttccagggggaaacgcctggtatctttatagtcctgtcgggtttcgccacctctgacttgagcgtcgatttttgtgatgctcgtcaggggggcggagcctatggaaaaacgccagcaacgcggcctttttacggttcctggccttttgctggccttttgctcacatgttctttcctgcgttatcccctgattctgtggataaccgtattaccgctagcatggatctcggggacgtctaactactaagcgagagtagggaactgccaggcatcaaataaaacgaaaggctcagtcggaagactgggcctttcgttttatctgttgtttgtcggtgaacgctctcctgagtaggacaaatccgccgggagcggatttgaacgttgtgaagcaacggcccggagggtggcgggcaggacgcccgccataaactgccaggcatcaaactaagcagaaggccatcctgacggatggcctttttgcgtttctacaaactcttcctgttagttagttacttaactagggataacagggtaatcggccccaaataatgattttattttgactgatagtgacctgttcgttgcaacaaattgataagcaatgcttttttataatgccaactttgtacaaaaaagcaggctttactgaagcttcggaattcggcgcgccagtgaattatcaactatgtataataaagttgaacgagaaacgtaaaatgatataaatatcaatatattaaattagattttgcataaaaaacagactacataatactgtaaaacacaacatatccagtcactatggcggcccgtgccatttcattacctctttctccgcacccgacatagatcgatcccacatatacctgccgttcactattatttagtgaaatgagatattatgatattttctgaattgtgattaaaaaggcaactttatgcccatgcaacagaaactataaaaaatacagagaatgaaaagaaacagatagattttttagttctttaggcccgtagtctgcaaatccttttatgattttctatcaaacaaaagaggaaaatagaccagttgcaatccaaacgagagtctaatagaatgaggtcgaaaagtaaatcgcgcgggtttgttactgataaagcaggcaagacctaaaatgtgtaaagggcaaagtgtatactttggcgtcaccccttacatattttaggtctttttttattgtgcgtaactaacttgccatcttcaaacaggagggctggaagaagcagaccgctaacacagtacataaaaaaggagacatgaacgatgaacatcaaaaagtttgcaaaacaagcaacagtattaacctttactaccgcactgctggcaggaggcgcaactcaagcgtttgcgaaagaaacgaaccaaaagccatataaggaaacatacggcatttcccatattacacgccatgatatgctgcaaatccctgaacagcaaaaaaatgaaaaatatcaagttcctgagttcgattcgtccacaattaaaaatatctcttctgcaaaaggcctggacgtttgggacagctggccattacaaaacgctgacggcactgtcgcaaactatcacggctaccacatcgtctttgcattagccggagatcctaaaaatgcggatgacacatcgatttacatgttctatcaaaaagtcggcgaaacttctattgacagctggaaaaacgctggccgcgtctttaaagacagcgacaaattcgatgcaaatgattctatcctaaaagaccaaaca
Vector information for pL34R12H-CmR-ccdB
Map of pL34R12H-CmR-ccdB
Location of Features
•Cm(R): 3709-69
•PI-SceI(R): 200-238
•ccdB: 573-878
•attR2: 919-1043(c)
•attL4: 1063-1158(c)
•I-SceI(R): 1177-1194(c)
•Gen(R): 1456-1989
•pUCorigin: 2082-2755
•rrnB T1: 2864-2907 (c)
•rrnB T2: 3039-3082 (c)
•I-SceI: 3107-3124
•attL3: 3143-3238
•attR1: 3271-3395
•PI-SceI: 3402-3440
(c) = complementary strand
Sequence of pL34R12H-CmR-ccdB
ggcttccatgtcggcagaatgcttaatgaattacaacagtactgcgatgagtggcagggcggggcgtaatttttttaaggcagttattggtgcgatcctctagctagaaattcgttcaagccgacgccgcttcgccggcgttaactcaagcgattagatgcactaagcacataattgctcacagccaaactatcaggtctgccatttcattacctctttctccgcacccgacatagatgatccggcttactaaaagccagataacagtatgcgtatttgcgcgctgatttttgcggtataagaatatatactgatatgtatacccgaagtatgtcaaaaagaggtgtgctatgaagcagcgtattacagtgacagttgacagcgacagctatcagttgctcaaggcatatatgatgtcaatatctccggtctggtaagcacaaccatgcagaatgaagcccgtcgtctgcgtgccgaacgctggaaagcggaaaatcaggaagggatggctgaggtcgcccggtttattgaaatgaacggctcttttgctgacgagaacagggactggtgaaatgcagtttaaggtttacacctataaaagagagagccgttatcgtctgtttgtggatgtacagagtgatattattgacacgcccgggcgacggatggtgatccccctggccagtgcacgtctgctgtcagataaagtctcccgtgaactttacccggtggtgcatatcggggatgaaagctggcgcatgatgaccaccgatatggccagtgtgccggtctccgttatcggggaagaagtggctgatctcagccaccgcgaaaatgacatcaaaaacgccattaacctgatgttctggggaatataaatgtcaggctcccttatacacagccagtctgcaggtcgaccatagtgactggatatgttgtgttttacagtattatgtagtctgttttttatgcaaaatctaatttaatatattgatatttatatcattttacgtttctcgttcagctttcttgtacaaagtggttcgataattccttaattaacaacttttctatacaaagttggcattataaaaaagcattgcttatcaatttgttgcaacgaacaggtcactatcagtcaaaataaaatcattatttggaatcgtcgagctgcacattaccctgttatccctaccatgagacgcacaccgtggaaacggatgaaggcacgaacccagttgacataagcctgttcggttcgtaaactgtaatgcaagtagcgtatgcgctcacgcaactggtccagaaccttgaccgaacgcagcggtggtaacggcgcagtggcggttttcatggcttgttatgactgtttttttgtacagtctatgcctcgggcatccaagcagcaagcgcgttacgccgtgggtcgatgtttgatgttatggagcagcaacgatgttacgcagcagcaacgatgttacgcagcagggcagtcgccctaaaacaaagttaggtggctcaagtatgggcatcattcgcacatgtaggctcggccctgaccaagtcaaatccatgcgggctgctcttgatcttttcggtcgtgagttcggagacgtagccacctactcccaacatcagccggactccgattacctcgggaacttgctccgtagtaagacattcatcgcgcttgctgccttcgaccaagaagcggttgttggcgctctcgcggcttacgttctgcccaggtttgagcagccgcgtagtgagatctatatctatgatctcgcagtctccggcgagcaccggaggcagggcattgccaccgcgctcatcaatctcctcaagcatgaggccaacgcgcttggtgcttatgtgatctacgtgcaagcagattacggtgacgatcccgcagtggctctctatacaaagttgggcatacgggaagaagtgatgcactttgatatcgacccaagtaccgccacctaacaattcgttcaagccgagatcggcttcccggccgcggagttgttcggtaaattgtcacaacgccgcccgccccgttccactgagcgtcagacccggtagaaaagatcaaaggatcttcttgagatcctttttttctgcgcgtaatctgctgcttgcaaacaaaaaaaccaccgctaccagcggtggtttgtttgccggatcaagagctaccaactctttttccgaaggtaactggcttcagcagagcgcagataccaaatactgttcttctagtgtagccgtagttaggccaccacttcaagaactctgtagcaccgcctacatacctcgctctgctaatcctgttaccagtggctgctgccagtggcgataagtcgtgtcttaccgggttggactcaagacgatagttaccggataaggcgcagcggtcgggctgaacggggggttcgtgcacacagcccagcttggagcgaacgacctacaccgaactgagatacctacagcgtgagctatgagaaagcgccacgcttcccgaagggagaaaggcggacaggtatccggtaagcggcagggtcggaacaggagagcgcacgagggagcttccagggggaaacgcctggtatctttatagtcctgtcgggtttcgccacctctgacttgagcgtcgatttttgtgatgctcgtcaggggggcggagcctatggaaaaacgccagcaacgcggcctttttacggttcctggccttttgctggccttttgctcacatgttctttcctgcgttatcccctgattctgtggataaccgtattaccgctagcatggatctcggggacgtctaactactaagcgagagtagggaactgccaggcatcaaataaaacgaaaggctcagtcggaagactgggcctttcgttttatctgttgtttgtcggtgaacgctctcctgagtaggacaaatccgccgggagcggatttgaacgttgtgaagcaacggcccggagggtggcgggcaggacgcccgccataaactgccaggcatcaaactaagcagaaggccatcctgacggatggcctttttgcgtttctacaaactcttcctgttagttagttacttaactagggataacagggtaatcggcccactagagattccaaataatgattttattttgactgatagtgacctgttcgttgcaacaaattgataagcaatgcttttttataatgccaactttgtataataaagttgaagcttcggaattcggcgcgccaagctatcaaacaagtttgtacaaaaaagctgaacgagaaacgtaaaatgatataaatatcaatatattaaattagattttgcataaaaaacagactacataatactgtaaaacacaacatatccagtcactatggcggccatctatgtcgggtgcggagaaagaggtaatgaaatggcaacataagcctcgttcggttcgtaagctgtaatgcaagtagcgtaactgccgtcacgcaactggtccagaaccttgaccgaacgcagcggtggtaacggcgcagtggcggttttcatggcttcttgttatgacatgtttttttggggtacagtctatgcctcgggcatccaagcagcaagcgcgttacgccgtgggtcgatgtttgatgttatggagcagcaacgatgttacgcagcagggcagtcgccctaaaacaaagttaaacatcatggagaaaaaaatcactggatataccaccgttgatatatcccaatggcatcgtaaagaacattttgaggcatttcagtcagttgctcaatgtacctataaccagaccgttcagctggatattacggcctttttaaagaccgtaaagaaaaataagcacaagttttatccggcctttattcacattcttgcccgcctgatgaatgctcatccggagttccgtatggcaatgaaagacggtgagctggtgatatgggatagtgttcacccttgttacaccgttttccatgagcaaactgaaacgttttcatcgctctggagtgaataccacgacgatttccggcagtttctacacatatattcgcaagatgtggcgtgttacggtgaaaacctggcctatttccctaaagggtttattgagaatatgtttttcgtctcagccaatccctgggtgagtttcaccagttttgatttaaacgtggccaatatggacaacttcttcgcccccgttttcacaatgggcaaatattatacgcaaggcgacaaggtgctgatgccgctggcgattcaggttcatcatgccgtttgtgat
Vector information for pGWTAC
Map of pGWTAC
Location of Features
•ccdB: 316-621
•attR2: 662-786 (c)
•loxP: 838-871
•I-SceI: 879-896
•RB:986-1010
•pRiA4replicon:1534-9417
•P1replicon: 10876-16590
•Kan(R): 9958-10776 (c)
•LB: 16593-16616
•attR1: 16679-16803
•Cm(R): 16916-17575
(c) = complementary strand
Sequence of pGWTAC
catcatccagccagaaagtgagggagccacggttgatgagagctttgttgtaggtggaccagttggtgattttgaacttttgctttgccacggaacggtctgcgttgtcgggaagatgcgtgatctgatccttcaactcagcaaaagttcgatttattcaacaaagccgccgtcccgtcaagtcagcgtaatgctctgccagtgttacaaccaattaaccaattctgattagaaaaactcatcgagcatcaaatgaaactgcaatttattcatatcaggattatcaataccatatttttgaaaaagccgtttctgtaatgaaggagaaaactcaccgaggcagttccataggatggcaagatcctggtatcggtctgcgattccgactcgtccaacatcaatacaacctattaatttcccctcgtcaaaaataaggttatcaagtgagaaatcaccatgagtgacgactgaatccggtgagaatggcaaaagcagcttatgcatttctttccagacttgttcaacaggccagccattacgctcgtcatcaaaatcactcgcatcaaccaaaccgttattcattcgtgattgcgcctgagcgagacgaaatacgcgatcgctgttaaaaggacaattacaaacaggaatcgaatgcaaccggcgcaggaacactgccagcgcatcaacaatattttcacctgaatcaggatattcttctaatacctggaatgctgttttcccggggatcgcagtggtgagtaaccatgcatcatcaggagtacggataaaatgcttgatggtcggaagaggcataaattccgtcagccagtttagtctgaccatctcatctgtaacatcattggcaacgctacctttgccatgtttcagaaacaactctggcgcatcgggcttcccatacaatcgatagattgtcgcacctgattgcccgacattatcgcgagcccatttatacccatataaatcagcatccatgttggaatttaatcgcggcctcgagcaagacgtttcccgttgaatatggctcataacaccccttgtattactgtttatgtaagcagacagttttattgttcatgatgatatatttttatcttgtgcaatgtaacatcagagattttgagacaccgcgcttgcggaagcatcagcaaataaggccagcacagccagcgcagttgccgctttggttcctgattctgttcttgatgaattaaacaaagcggcacagtaacaaaggacttcattgataatttttcttcaggaggaagacatgtcattcttttctacgttaaaaacagctttgtctttgaaggagaaacttgctgctactggtgttcttgttctgatttgcgcacttgttggtgctgggtttgcatgggaacgtcatcagctaaagcaagccatagagaaaattggcagtcttgatcaggctgttaaggaacgtgataagtcaataatggatcttaaccagaccattgagacgatgaacaaagcagagcaacattttcacagccaggaagtgaaaaatgaatcagaacaagccaagtatgctgacaggcaaatggaacgaaaagctgaagttcagaaacaactggttgcggcgggtaatgttcgccagcgtattcctgctgacactcagcggttgctccgggagtcgatcagcgaatttaacgccgacgccgacaaaggttaaccaccctgcccccaaaagtgcatttatgtgtaggatgccagagtttagcagtgaatattttgatgatctgccagcgtatatcctcgatacagaaacgatgctgatggggattaacaggaagaatcgcaacgttaatgattacaaccgagctattagcggtaactaaaagggatttttatgtctgataaagtaacagtaaagcaaactatcaacaaagcgacttcaatctacaaaattgagcaaatcactgttggcaagccaggatctgaacaataccgtcgtgctttcgagcttgccgatcagcttggtttaaaacacccggattgcattgagcatgtatttccgacctatgctgatgagcaatgtactcatgttcttaccgaagaggattttttcagcactgaagaacgagaaggcgttgatcgctgcattggtgtgatttgttcttcggtaagtgatgagttattccctaatgtgcctgaatatggtggtattggataccaattcctgtacgagggcgatgagcttaaatgctatgaacatggtcttctcatcgaaagcgtagaataatacgactcccttccaaccggctacgttggccggtttttcacttatccacattatccactggatagatccaataatcaggtccatacagatcccaattagatccatatagatccctgatcgttgcaggccgcgccacgtctggcttagaagtgtatcgcgatgtgtgctggagggaaaacgatgtgtgctggagggataaaaatgtgtgctgacgggttgctaatgtgtgctggcgggatataggatgtgtgttgacgggaaagctagcttgggtagttatcaccacttataaaaactatccacacaattcggaaaaagtaatatgaatcaatcatttatctccgatattctttacgcagacattgaaagtaaggcaaaagaactaacagttaattcaaacaacactgtgcagcctgtagcgttgatgcgcttgggggtattcgtgccgaagccatcaaagagcaaaggagaaagtaaagagattgatgccaccaaagcgttttcccagctggagatagctaaagccgagggttacgatgatattaaaatcaccggtcctcgactcgatatggatactgatttcaaaacgtggatcggtgtcatctacgcgttcagcaaatacggcttgtcctcaaacaccatccagttatcgtttcaggaattcgctaaagcctgtggtttcccctcaaaacgtctggatgcgaaactgcgtttaaccattcatgaatcacttggacgcttgcgtaacaagggtatcgcttttaagcgcggaaaagatgctaaaggcggctatcagactggtctgctgaaggtcgggcgttttgatgctgaccttgatctgatagagctggaggctgattcgaagttgtgggagctgttccagcttgattatcgcgttctgttgcaacaccacgccttgcgtgcccttccgaagaaagaagctgcacaagccatttacactttcatcgaaagccttccgcagaacccgttgccgctatcgttcgcgcgaatccgtgagcgcctggctttgcagtcagctgttggcgagcaaaaccgtatcattaagaaagcgatagaacagcttaaaacaatcggctatctcgactgttctattgagaagaaaggccgggaaagttttgtaatcgtccattctcgcaatccaaagctgaaactccccgaataagtgtgtgctggagggaaaccgcattaaaaagatgtgtgctgccgggaaggcttgtccaatttcctgtttttgatgtgcgctggagggggacgcccctcagtttgcccagactttccctccagcacacatctgtccatccgcttttccctccagcacacatcgaagctgccgggcaagccgttctcaccagttgatagagagtgaagctagcttggctgcccattgaagcaggaaatcaccaaaatgattcaggctacaacctgaacgtagaagaaatccgcgtcctttatgcgtggaggatgccaaagcatgttgtgacacacttggcaaaggagtaagcatgcagagaatgctatgtacaagcatctacgcatacattattattttatgcagcatttttaattaaattcaaaaatacagcataaaggatgactttcgatgagtgattccagccagcttcacaaggttgctcaaagagcaaacagaatgctcaatgttctgactgaacaagtacagttgcaaaaggatgagctacacgcgaacgagttttaccaggtctatgcgaaagcggcactggcaaaattgcctctactgactcgagcgaacgttgactatgccgtaagtgaaatggaagaaaagggttatgttttcgataaacgccctgctggctcttcaatgaaatatgcgatgtcaattcagaacatcattgacatatatgaacatcgcggagtgccaaaataccgggatcgctacagcgaagcgtatgtgattttcatctccaatcttaaaggcggtgtgtcaaaaactgtatcgacggtttctctggcgcatgcaatgcgtgctcaccctcatcttcttatggaggatttaaggattctggttattgaccttgatccgcaatcttcagcaacgatgtttttaagccataaacactctattggtatcgtaaacgcaacatctgcacaggctatgttgcagaatgtaagccgtgaagagctgttagaggagtttattgttccttctgttgtacctggggttgacgttatgcctgcgtcgattgacgatgcctttattgcatccgattggagagagctgtgcaatgagcatctaccgggtcagaacatccatgctgtcctgaaagaaaatgtgattgataagctgaagagcgattatgactttatcctcgttgatagtggtcctcaccttgacgccttcctgaaaaatgctttggcctcggccaatatactgtttacacctctgccgccagcaactgtcgatttccactcatcgcttaaatacgttgcccgccttcctgagttggtgaaactcatttcggatgaaggctgcgagtgccagcttgcgactaacattggttttatgtccaagttgagtaacaaggcagaccataagtattgccatagcctggctaaagaagtgttcggtggggatatgcttgatgtcttcctccctcgccttgacggttttgaacgctgcggcgagtcttttgacactgttatttcagctaacccggcaacgtatgttggtagtgctgatgcattgaagaacgcgcgaattgccgcggaagattttgctaaagcagtttttgaccgtattgaatttatcagatctaactgaggagtaagaaacccccatgtcaaagaaaaacagaccaacaattgggcgaacccttaatccttcaatattaagcggatttgatagttcttcagcctctggcgatcgagtcgagcaggtattcaagttatcaactggtcgccaggccacatttattgaagaggtaatacctccgaaccaggtagaaagcgatacctttgttgatcagcataacaacgggcgtgaccaggcatctcttacgccaaaatcattaaaaagtatccgaagcactattaagcatcagcaattttaccctgcaataggtgttagacgggctacagggaaaattgaaattttggatggttcccggcgtcgagcttctgccatcttagagaacgtagggttgcgggttttagtcacggaccaggagatcagcgttcaggaagcgcaaaatttagcgaaagacgttcagacagcattgcagcacagcattcgagaaataggtctgcgtttgatgcgaatgaaaaatgatgggatgagtcagaaggatattgcagccaaagaagggctgtctcaggcgaaggtcacgcgtgctctccaggcagcgagtgctccggaagaattagtcgcccttttccctgtgcagtcggaattaaccttttcggactacaaaacgctttgtgctgttggcgacgaaatggggaacaagaatttagagtttgatcagcttattcaaaacatatccccggaaataaacgacatcttatccattgaagaaatggccgaagatgaagttaaaaataaaatcctgcgcttgataacaaaggaagcctcactactcacggataaaggttctaaagataagtccgtagttactgaattatggaaatttgaggacaaggatcgctttgcaaggaagcgcgtgaaaggccgtgcattttcttatgagtttaatcgactctcaaaagagttacaggaagaactcgacaggatgattgggcatatccttagaaagagcctcgataaaaagccgaagccttaaactttcgccattcaaatttcactattaactgactgtttttaaagtaaattactctaaaatttcaaggtgaaatcgccacgatttcaccttggattttaccttcctcccctcctcccgaaaaaaataaaaaaattgcttgtcacgagaaagtcaacaagtgactttcaataaaatctcttccgaaaagggattcacacaagtgccttgtgtttaaggaagagtaaattgagtaacttacgcgaataccagaatcgtattgcagatatcgcaaaacgctctaaagctgtgcttggctgggcaagcactgcgcagttcggtactgataaccaattcattaaagatgatgccgcgcgtgccgcatctatccttgaagctgcacgtaaagacccggtttttgcgggtatctctgataatgccaccgctcaaatcgctacagcgtgggcaagtgcactggctgactacgccgcagcacataaatctatgccgcgtccggaaattctggcctcctgccaccagacgctggaaaactgcctgatagagtccacccgcaatagcatggatgccactaataaagcgatgctggaatctgtcgcagcagagatgatgagcgtttctgacggtgttatgcgtctgcctttattcctcgcgatgatcctgcctgttcagttgggggcagctaccgctgatgcgtgtaccttcattccggttacgcgtgaccagtccgacatctatgaagtctttaacgtggcaggatatattgtggtgtaaacaggatccaagcttcgaccatcctccaaaatcggatctggttccgcgtccatggtcgaatcaaacaagtttgtacaaaaaagctgaacgagaaacgtaaaatgatataaatatcaatatattaaattagattttgcataaaaaacagactacataatactgtaaaacacaacatatccagtcactatggcggccggccgcattaggcaccccaggctttacactttatgcttccggctcgtataatgtgtggattttgagttaggatccgtcgagattttcaggagctaaggaagctaaaatggagaaaaaaatcactggatataccaccgttgatatatcccaatggcatcgtaaagaacattttgaggcatttcagtcagttgctcaatgtacctataaccagaccgttcagctggatattacggcctttttaaagaccgtaaagaaaaataagcacaagttttatccggcctttattcacattcttgcccgcctgatgaatgctcatccggaattccgtatggcaatgaaagacggtgagctggtgatatgggatagtgttcacccttgttacaccgttttccatgagcaaactgaaacgttttcatcgctctggagtgaataccacgacgatttccggcagtttctacacatatattcgcaagatgtggcgtgttacggtgaaaacctggcctatttccctaaagggtttattgagaatatgtttttcgtctcagccaatccctgggtgagtttcaccagttttgatttaaacgtggccaatatggacaacttcttcgcccccgttttcaccatgggcaaatattatacgcaaggcgacaaggtgctgatgccgctggcgattcaggttcatcatgccgtctgtgatggcttccatgtcggcagaatgcttaatgaattacaacagtactgcgatgagtggcagggcggggcgtaatctagaggatccggcttactaaaagccagataacagtatgcgtatttgcgcgctgatttttgcggtataagaatatatactgatatgtatacccgaagtatgtcaaaaagaggtgtgctatgaagcagcgtattacagtgacagttgacagcgacagctatcagttgctcaaggcatatatgatgtcaatatctccggtctggtaagcacaaccatgcagaatgaagcccgtcgtctgcgtgccgaacgctggaaagcggaaaatcaggaagggatggctgaggtcgcccggtttattgaaatgaacggctcttttgctgacgagaacagggactggtgaaatgcagtttaaggtttacacctataaaagagagagccgttatcgtctgtttgtggatgtacagagtgatattattgacacgcccgggcgacggatggtgatccccctggccagtgcacgtctgctgtcagataaagtctcccgtgaactttacccggtggtgcatatcggggatgaaagctggcgcatgatgaccaccgatatggccagtgtgccggtctccgttatcggggaagaagtggctgatctcagccaccgcgaaaatgacatcaaaaacgccattaacctgatgttctggggaatataaatgtcaggctcccttatacacagccagtctgcaggtcgaccatagtgactggatatgttgtgttttacagtattatgtagtctgttttttatgcaaaatctaatttaatatattgatatttatatcattttacgtttctcgttcagctttcttgtacaaagtggtttgattcgacccgggatccggctgctaacaaagcccgaaaggagcggccgcataacttcgtatagcatacattatacgaagttatgggccgcattaccctgttatccctaggccgcaattaggcctacccactagacccttaattctccgctcatgatcagattgtcgtttcccgccttcagtttaaactatcagtgtttgacaggatatattggcgggtaaacctaagagaaaagagcgtttattagaataatcggatatttaaaagggcgtgaaaaggtttatccgttcgtccatttgtatgtgcatgccaaccacagggttcccctcgggagtgcttggcattccgtgcgataatgacttctgttcaaccacccaaacgtcggaaagcctgacgacggagcagcattccaaaaagatcccttggctcgtctgggtcggctagaaggtcgagtgggctgctgtggcttgatccctcaacgcggtcgcggacgtagcgcagcgccgaaaaatcctcgatcgcaaatccgacgctgtcgaaaagcgtgatctgcttgtcgctctttcggccgacgtcctggccagtcatcacgcgccaaagttccgtcacaggatgatctggcgcgagttgctggatctcgccttcaatccgggcctgtggcgggaactccacgaaaatatccgaacgcagcaagatcgtcgatcctccggcgttcagcctgtgccacagccgacaggatggtgaccgaatcctacaaggtagaatccgcctgagtcgcaagggtgacttcgcctatattggacgacggcgcgcagagggcgacctctttttgggttacgattgtaggattatcactaaaacaatacatgaacatattcaaatggcaatctctctaaggcattggaaataaatacaaataacagttgggtggagtttttcgacctgagggcgttaaccttctgttaacctaaaagctcttgcccaaacagcagaatcggcgctaattgccagcggcggaacttttccagtttcgcgaaaagtatcgccactggcaaggaatgggtttgagatggcgaagtctgtcctaaaagcagcgcctgtagttgtagggttgacggccttgatggagcgtcatgccgatgccctctcgagccaacttcaagcacatcatcttaaggttttcccgccgcattccgagaagggcattcgaacattcgggccatcggaggcgtccaagctgctcggcgttggcgagtcatatttacggcagaccgcgtctgagatgccagagttgaatgttagcatgagcccgggtggcaggcgaatgttctcaattgaaga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