hhhhhhhhhhhhh Solution Golden Gate

June 30, 2025, 6:38 p.m.

  • 4 Parts 8 Primers
  • 60.62C Average Primer Tm 31nt Average Primer 3 BLAST Parts 1 Synthetic Parts 707nt Longest Part 111nt Shortest Part
  • 94.00% BLAST nt 6.00% Synthetic nt
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Parts Download Parts

Below is a summary table of the DNA sequences that make up this assembly solution. Further details on each part can be viewed by clicking View Part for each listed below this table.

Query Start/End: The start/end index of the part on the insert sequence. Defaults to 0.
Subject Start/End: The start/end index of the part on its original sequence. Defaults to 0.
Position: The location of the part in the assembly solution.
Name Database Length Extended Length Query Start Query End Subject Start Subject End Position
120714-addgene addgene 117 136 4 121 21069 20937 0
74625-addgene addgene 707 741 125 832 2063 2773 1
Synthetic-1.112 NONE 111 133 836 947 0 0 2
71655-addgene addgene 704 723 951 1655 6958 7674 3
pBR322_b NONE 4360 4391 1666 6026 0 0 4

Plasmid Map

Parts List

120714-addgene addgene View Part

136nt Extended 117nt Template

  • Assembly Sequence: GGTCTCNGACAGgtgagctgatacc...gcgttggccgattcattaNGAGACC
  • Template Sequence: gtgagctgataccgctcgccgcagc...tccccgcgcgttggccgattcatta

74625-addgene addgene View Part

741nt Extended 707nt Template

  • Assembly Sequence: GGTCTCNattaatgcagctggcacg...catggacgagctgtacaaNGAGACC
  • Template Sequence: atggtgagcaagggcgaggacgaca...ccggcggcatggacgagctgtacaa

Synthetic-1.112 View Part

133nt Extended 111nt Template

  • Assembly Sequence: GGTCTCNacaagtaaggtgatacca...gtcccaattcttgttgaaNGAGACC
  • Template Sequence: ggtgataccagcatcgtcttgatgc...tggagttgtcccaattcttgttgaa

71655-addgene addgene View Part

723nt Extended 704nt Template

  • Assembly Sequence: GGTCTCNtgaatatgagtaaaggag...tacacatggcatggatgaNGAGACC
  • Template Sequence: atgagtaaaggagaagaacttttca...ctgggattacacatggcatggatga

pBR322_b View Part

4391nt Extended 4360nt Template Internal Cutsite(s) - Adjust probability of success for future experiments

  • Assembly Sequence: GGTCTCNatgagctctacaaataaC...GAATTCTCATGTTTGACANGAGACC
  • Template Sequence: CTTATCATCGATAAGCTTTAATGCG...TCTTCAAGAATTCTCATGTTTGACA

Primers Download Primers

Below is a summary table of the primers needed for each part of this assembly solution (shown above). Further details on each primer can be viewed by clicking View Primer for each listed below this table.

hp Tm (C): The melting temperature of the primer's hairpin (hp) structure.
hp ΔG: The Gibbs free energy of the primer's hairpin (hp) structure.
hd Tm (C): The melting temperature of the primer's homodimer (hd) structure.
hd ΔG: The Gibbs free energy of the primer's homodimer (hd) structure.
Name Type Tm (C) GC% Hairpin hp Tm (C) hp ΔG Homodimer hd Tm (C) hd ΔG
120714-addgene forward primer fwd 60.471 32.14 True 41.615 -385.626 True 19.277 -7324.651
120714-addgene reverse primer rvs 60.314 33.33 True 46.105 -547.579 True 2.079 -5455.321
74625-addgene forward primer fwd 62.012 45.24 True 58.993 -1443.514 True 16.903 -7044.691
74625-addgene reverse primer rvs 61.541 36.0 False 0.000 0.000 True -8.830 -3413.286
Synthetic-1.112 forward primer fwd 59.473 35.48 True 41.613 -271.143 True 0.084 -3163.304
Synthetic-1.112 reverse primer rvs 59.592 26.92 True 53.877 -1047.648 True -23.073 -2736.010
71655-addgene forward primer fwd 60.173 22.86 True 37.663 -70.162 True -20.930 -3085.011
71655-addgene reverse primer rvs 60.961 33.33 False 0.000 0.000 True 3.340 -4081.791
pBR322_b forward primer fwd 60.806 10.42 True 43.905 -1526.673 True 15.976 -6721.761
pBR322_b reverse primer rvs 60.872 0.0 True 33.558 399.571 True 9.560 -5625.021

Primers List

120714-addgene forward primer View Primer

fwd 60.471°C Hairpin Homodimer

  • Sequence: GGTCTCNGACAGgtgagctgataccgct

120714-addgene reverse primer View Primer

rvs 60.314°C Hairpin Homodimer

  • Sequence: GGTCTCNtaatgaatcggccaacg

74625-addgene forward primer View Primer

fwd 62.012°C Hairpin Homodimer

  • Sequence: GGTCTCNattaatgcagctggcacgacatggtgagcaagggc

74625-addgene reverse primer View Primer

rvs 61.541°C Homodimer

  • Sequence: GGTCTCNttgtacagctcgtccatg

Synthetic-1.112 forward primer View Primer

fwd 59.473°C Hairpin Homodimer

  • Sequence: GGTCTCNacaagtaaggtgataccagcatcg

Synthetic-1.112 reverse primer View Primer

rvs 59.592°C Hairpin Homodimer

  • Sequence: GGTCTCNttcaacaagaattgggaca

71655-addgene forward primer View Primer

fwd 60.173°C Hairpin Homodimer

  • Sequence: GGTCTCNtgaatatgagtaaaggagaagaactttt

71655-addgene reverse primer View Primer

rvs 60.961°C Homodimer

  • Sequence: GGTCTCNtcatccatgccatgtgt

pBR322_b forward primer View Primer

fwd 60.806°C Hairpin Homodimer

  • Sequence: GGTCTCNatgagctctacaaataaCTTATCATCGATAAGCTTTAATGC

pBR322_b reverse primer View Primer

rvs 60.872°C Hairpin Homodimer

  • Sequence: GGTCTCNTGTCAAACATGAGAATTCTTGAAG
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