Skip to main content

Table 3 Primers used for targeted gene deletions

From: Trehalose synthesis in Aspergillus niger: characterization of six homologous genes, all with conserved orthologs in related species

Primer name

Sequence 5′-3′

Purpose

pyrGN2

CACATGCCTCATTTTGACCA

Mutant confirmation

PyrtpsAup

ACCGTTGGAAGGTGGGATCCTATGGATCTCAGAA

Amplifies pyrG with 3' tpsA overhangs

PyrtpsAdown

CCTTTCAGAATGAGTGTGAGCGGATAACAATTTC

tpsAup

CCATCTGTCTAGCTCTTCATCCCC

tpsA, upstream fragment

tpsApyrup

GATCCATAGGATCCCACCTTCCAACGGTGTAGAGACTCC

tpsApyrdown

TTATCCGCTCACACTCATTCTGAAAGGTGGGGTTTTC

tpsA, downstream fragment

tpsAdown

GCAAGATTCCCGCATCCATC

tpsAupN1

CAACCCCACCAGTTCTCTCAAG

Amplification of KO-fragment

tpsAdownN1

AAAGGGAGTTCCAAGCAGCCTG

pyrtpsBup*

ATCTGCTCTGCCTGGGATCCTATGGATCTCAGAA

Amplifies pyrG with 3′ tpsB overhangs

pyrtpsBdown

CTGCCCATCACCATGTGAGCGGATAACAATTTC

tpsBup*

TTGAACCCTTGAAACCGAACAC

tpsB, upstream fragment

tpsBpyrGup*

GATCCATAGGATCCCAGGCAGAGCAGATACTTACCCGTC

tpsBpyrGdown

TTATCCGCTCACATGGTGATGGGCAGACGATTG

tpsB, downstream fragment

tpsBdown

TGCTAAAGAGGGTGTGGGATTG

tpsBupN3

TCCCGATTGGTAGAATCCCTAAAG

Amplification of tpsB KO-fragment

tpsBdownN3

CATGCGAAAATGACAGGAACATTC

pyrGuphind

TAAAAGCTTCTATATTGATCCTTA

pyrG, KO of tpsC

pyrGdown

TGTGAGCGGATAACAATTTC

tpsCupN-2

TGCCGAATTGACGTGCGTAGAG

Cloning of tpsC

tpsCdownN-2

TGGTGGTGAACCTTTCGTTGTTC

tpsCupN5

CCCTCCATACTTACTCCATACATCTCG

Amplification of tpsC KO-fragment

tpsCdownN5

CCAGCTTGACACATCCAACATAAC

pyrtppAup

CCTGTCCCCGCTTCAAGAAAGGGATCCTATGGATCTCAGAA

pyrG with 3′ tppA overhangs

pyrtppAdown

GAGTCATCAGTGCTGCTTTCTGCTGTGAGCGGATAACAATTTC

TppAup

TGTTGGAAGCGTCTTTCTGCC

tppA, upstream fragment

tppApyrup

TTCTGAGATCCATAGGATCCCTTTCTTGAAGCGGGGACAGG

tppApyrdown

GAAATTGTTATCCGCTCACAGCAGAAAGCAGCACTGATGACTC

tppA, downstream fragment

tppAdown

TGTCCGATTGGGGGTGATTG

tppAupN1

TGAGGAGGCGTTGTCAAAAGATAG

Amplification of tppA KO-fragment

tppAdownN1

CGATTGGGGGTGATTGGCTTAC

pyrtppBup

CGGTAGGTTAGGGATCCTATGGATCTCAGAA

Amplification of A. oryzae pyrG with 3′ tppB overhangs

pyrtppBdown

GTTTGTCTTGTGTGAGCGGATAACAATTTC

tppBup

ATACCAAGCAATCGCCCAAGCCAG

tppB, upstream fragment

tppBpyrGup

TCCATAGGATCCCTAACCTACCGCCCAAAGAGAGAGC

tppBpyrGdown

TTGTTATCCGCTCACACAAGACAAACGATGCGGAATG

tppB, downstream fragment

tppBdown

CGTATCCTGGACTTTCAGCACG

tppBupN1

TTTTCGACCTTGGTGGGTGCTTCC

Amplification of tppB KO-fragment

tppBdownN1

GAGACATTGTCGGTCAGTGAGGTAG

pyrtppCup

TGTCCTTCAGGGATCCTATGGATCTCAGAA

pyrG with 3′ tppC overhangs

pyrtppCdown*

CTGTTCAGCATTGTGAGCGGATAACAATTTC

tppCup

ATGAGGTGATAGTCGTGGACCCAG

tppC, upstream fragment

tppCpyrup

TCCATAGGATCCCTGAAGGACAAAGACAGGCTGAAG

tppCpyrdown*

TTGTTATCCGCTCACAATGCTGAACAGATGATCCCCAG

tppC, downstream fragment

tppCdown*

TCGAGGTAGAGGTTCCCTTTCG

tppCupN1*

CGATAGTCTTTGCGAACAGACGG

Amplification of tppC KO-fragment

tppCdownN1*

CGAGGTAGAGGTTCCCTTTCGATG

tpsBupN1

CCCTTTCCCGATTGGTAGAATC

Amplification of tpsB/tppC double mutant

  1. *Also used in the design of the tpsB/tppC double mutant.