scarless test Solution Golden Gate

June 30, 2025, 6:05 p.m.

  • 4 Parts 8 Primers
  • 60.42C Average Primer Tm 34nt Average Primer 3 BLAST Parts 1 Synthetic Parts 698nt Longest Part 111nt Shortest Part
  • 90.00% BLAST nt 10.00% 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
116061-addgene addgene 117 156 4 121 7199 7067 0
64128-addgene addgene 698 732 125 823 5036 5746 1
Synthetic-1.112 NONE 111 142 827 938 0 0 2
35363-addgene addgene 693 712 942 1635 5726 6442 3
pBR322_b NONE 4340 4382 1675 6035 0 0 4

Plasmid Map

Parts List

116061-addgene addgene View Part

156nt Extended 117nt Template

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

64128-addgene addgene View Part

732nt Extended 698nt Template

  • Assembly Sequence: GGTCTCNattaatgcagctggcacg...caccggcggcatggacgaNGAGACC
  • Template Sequence: atggtgagcaagggcgaggaggata...gccactccaccggcggcatggacga

Synthetic-1.112 View Part

142nt Extended 111nt Template

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

35363-addgene addgene View Part

712nt Extended 693nt Template

  • Assembly Sequence: GGTCTCNtgaatatgagtaaaggag...gctgctgggattacacatNGAGACC
  • Template Sequence: atgagtaaaggagaagaacttttca...tgtaactgctgctgggattacacat

pBR322_b View Part

4382nt Extended 4340nt Template Internal Cutsite(s) - Adjust probability of success for future experiments

  • Assembly Sequence: GGTCTCNacatggcatggatgagct...CGAGGCCCTTTCGTCTTCNGAGACC
  • Template Sequence: CTTATCATCGATAAGCTTTAATGCG...CGTATCACGAGGCCCTTTCGTCTTC

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
116061-addgene forward primer fwd 60.471 18.75 True 41.615 -385.626 True 19.277 -7324.651
116061-addgene reverse primer rvs 60.314 33.33 True 46.105 -547.579 True 2.079 -5455.321
64128-addgene forward primer fwd 62.012 45.24 True 58.993 -1443.514 True 16.903 -7044.691
64128-addgene reverse primer rvs 60.583 45.0 False 0.000 0.000 True -27.504 -2497.791
Synthetic-1.112 forward primer fwd 59.473 40.0 True 37.584 -92.277 True 18.599 -1553.373
Synthetic-1.112 reverse primer rvs 59.592 26.92 True 53.877 -1047.648 True -23.073 -2736.010
35363-addgene forward primer fwd 60.173 22.86 True 37.663 -70.162 True -20.930 -3085.011
35363-addgene reverse primer rvs 59.208 33.33 False 0.000 0.000 True -33.069 -2810.310
pBR322_b forward primer fwd 60.806 18.64 True 42.695 -1074.653 True 29.241 -9147.601
pBR322_b reverse primer rvs 61.570 0.0 True 50.507 -1289.519 True -4.088 -5260.081

Primers List

116061-addgene forward primer View Primer

fwd 60.471°C Hairpin Homodimer

  • Sequence: GGTCTCNCTTCAAGAATTCTCATGTTTGACAGgtgagctgataccgct

116061-addgene reverse primer View Primer

rvs 60.314°C Hairpin Homodimer

  • Sequence: GGTCTCNtaatgaatcggccaacg

64128-addgene forward primer View Primer

fwd 62.012°C Hairpin Homodimer

  • Sequence: GGTCTCNattaatgcagctggcacgacatggtgagcaagggc

64128-addgene reverse primer View Primer

rvs 60.583°C Homodimer

  • Sequence: GGTCTCNtcgtccatgccgc

Synthetic-1.112 forward primer View Primer

fwd 59.473°C Hairpin Homodimer

  • Sequence: GGTCTCNacgagctgtacaagtaaggtgataccagcatcg

Synthetic-1.112 reverse primer View Primer

rvs 59.592°C Hairpin Homodimer

  • Sequence: GGTCTCNttcaacaagaattgggaca

35363-addgene forward primer View Primer

fwd 60.173°C Hairpin Homodimer

  • Sequence: GGTCTCNtgaatatgagtaaaggagaagaactttt

35363-addgene reverse primer View Primer

rvs 59.208°C Homodimer

  • Sequence: GGTCTCNatgtgtaatcccagcag

pBR322_b forward primer View Primer

fwd 60.806°C Hairpin Homodimer

  • Sequence: GGTCTCNacatggcatggatgagctctacaaataaCTTATCATCGATAAGCTTTAATGC

pBR322_b reverse primer View Primer

rvs 61.570°C Hairpin Homodimer

  • Sequence: GGTCTCNGAAGACGAAAGGGCCT
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