pMJ506: pLM005-Cre17.g724300-Venus-3xFLAG-3xCtermSAGA2

Description: PSAK (Cre17.g724300) gDNA coding sequence, spanning the start codon to the triplet just before the stop codon, had been previously cloned in frame into pLM005 by Gibson assembly (as described in https://doi.org/10.1073/pnas.1522866113; pMJ280  = pLM005-Cre17.g724300-Venus-3xFLAG). That plasmid was then reengineered by restriction digestion and T4 ligation to add in-frame immediately after the 3xFLAG a synthetic fragment containing three copies of the sequence coding for the 15 C-terminal amino acids of SAGA2 (Cre 09.g394621), interspersed with a short flexible linker (GGGGGS)2.

Selection in E. coli: ampicillin
Selection in C. reinhardtii: paromomycin
Host strain: DH5α
Transformed into C. reinhardtii strain cMJ030 with carrier DNA to produce CC-5651

Plasmid: 8275 bp DNA circular
CDS	0784..1587	“APHVIII"
CDS	2699..4018	“PSAK”
CDS	4019..5098	“VENUS-3xFLAG-3xSAGA2Cterm fusion protein”
TRANSLATION: NGDLGGSGGRMVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDGNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLLEFVTAAGITLGMDELIKGGGDYKDHDGDYKDHDIDYKDDDDKGGGGSGGGGSPVRRTAIPANWRDALGGGGSGGGGSGGGGSPVRRTAIPANWRDALGGGGSGGGGSGGGGSPVRRTAIPANWRDAL-

Plasmid sequence	1..8275
TCGCGCGTTTCGGTGATGACGGTGAAAACCTCTGACACATGCAGCTCCCGGAGACGGTCACAGCTTGTCTGTAAGCGGATGCCGGGAGCAGACAAGATATCCTTGACATGATTGGTGCGTATGTTTGTATGAAGCTACAGGACTGATTTGGCGGGCTATGAGGGCGGGGGAAGCTCTGGAAGGGCCGCGATGGGGCGCGCGGCGTCCAGAAGGCGCCATACGGCCCGCTGGCGGCACCCATCCGGTATAAAAGCCCGCGACCCCGAACGGTGACCTCCACTTTCAGCGACAAACGAGCACTTATACATACGCGACTATTCTGCCGCTATACATAACCACTCAGCTAGCTTAAGATCCCATCAAGCTTGCATGCCGGGCGCGCCAGAAGGAGCGCAGCCAAACCAGGATGATGTTTGATGGGGTATTTGAGCACTTGCAACCCTTATCCGGAAGCCCCCTGGCCCACAAAGGCTAGGCGCCAATGCAAGCAGTTCGCATGCAGCCCCTGGAGCGGTGCCCTCCTGATAAACCGGCCAGGGGGCCTATGTTCTTTACTTTTTTACAAGAGAAGTCACTCAACATCTTAAAATGGCCAGGTGAGTCGACGAGCAAGCCCGGCGGATCAGGCAGCGTGCTTGCAGATTTGACTTGCAACGCCCGCATTGTGTCGACGAAGGCTTTTGGCTCCTCTGTCGCTGTCTCAAGCAGCATCTAACCCTGCGTCGCCGTTTCCATTTGCAGGATGGCCACTCCGCCCTCCCCGGTGCTGAAGAATTTCGAAGCATGGACGATGCGTTGCGTGCACTGCGGGGTCGGTATCCCGGTTGTGAGTGGGTTGTTGTGGAGGATGGGGCCTCGGGGGCTGGTGTTTATCGGCTTCGGGGTGGTGGGCGGGAGTTGTTTGTCAAGGTGGCAGCTCTGGGGGCCGGGGTGGGCTTGTTGGGTGAGGCTGAGCGGCTGGTGTGGTTGGCGGAGGTGGGGATTCCCGTACCTCGTGTTGTGGAGGGTGGTGGGGACGAGAGGGTCGCCTGGTTGGTCACCGAAGCGGTTCCGGGGCGTCCGGCCAGTGCGCGGTGGCCGCGGGAGCAGCGGCTGGACGTGGCGGTGGCGCTCGCGGGGCTCGCTCGTTCGCTGCACGCGCTGGACTGGGAGCGGTGTCCGTTCGATCGCAGTCTCGCGGTGACGGTGCCGCAGGCGGCCCGTGCTGTCGCTGAAGGGAGCGTCGACTTGGAGGATCTGGACGAGGAGCGGAAGGGGTGGTCGGGGGAGCGGCTTCTCGCCGAGCTGGAGCGGACTCGGCCTGCGGACGAGGATCTGGCGGTTTGCCACGGTGACCTGTGCCCGGACAACGTGCTGCTCGACCCTCGTACCTGCGAGGTGACCGGGCTGATCGACGTGGGGCGGGTCGGCCGTGCGGACCGGCACTCCGATCTCGCGCTGGTGCTGCGCGAGCTGGCCCACGAGGAGGACCCGTGGTTCGGGCCGGAGTGTTCCGCGGCGTTCCTGCGGGAGTACGGGCGCGGGTGGGATGGGGCGGTATCGGAGGAAAAGCTGGCGTTTTACCGGCTGTTGGACGAGTTCTTCTGAGGGACCTGATGGTGTTGGTGGCTGGGTAGGGTTGCGTCGCGTGGGTGACAGCACAGTGTGGACGTTGGGATCCCCGCTCCGTGTAAATGGAGGCGCTCGTTGATCTGAGCCTTGCCCCCTGACGAACGGCGGTGGATGGAAGATACTGCTCTCAAGTGCTGAAGCGGTAGCTTAGCTCCCCGTTTCGTGCTGATCAGTCTTTTTCAACACGTAAAAAGCGGAGGAGTTTTGCAATTTTGTTGGTTGTAACGATCCTCCGTTGATTTTGGCCTCTTTCTCCATGGGCGGGCTGGGCGTATTTGAAGCGGGTACCAGCTTTTGTTCCCTTTAGTGAGGGTTAATTGCGACCAATCGTCACACGAGCCCTCGTCAGAAACACGTCTCCGCCACGCTCTCCCTCTCACGGCCGACCCCGCAGCCCTTTTGCCCTTTCCTAGGCCACCGACAGGACCCAGGCGCTCTCAGCATGCCTCAACAACCCGTACTCGTGCCAGCGGTGCCCTTGTGCTGGTGATCGCTTGGAAGCGCATGCGAAGACGAAGGGGCGGAGCAGGCGGCCTGGCTGTTCGAAGGGCTCGCCGCCAGTTCGGGTGCCTTTCTCCACGCGCGCCTCCACACCTACCGATGCGTGAAGGCAGGCAAATGCTCATGTTTGCCCGAACTCGGTGTCCTTAAAAAGCCGCTTCTTGTCGTCGTTCCGAGACATGTTAGCAGATCGCAGTGCCACCTTTCCTGACGCGCTCGGCCCCATATTCGGACGCAATTGTCATTTGTAGCACAATTGGAGCAAATCTGGCGAGGCAGTAGGCTTTTAAGTTGCAAGGCGAGAGAGCAAAGTGGGACGCGGCGTGATTATTGGTATTTACGCGACGGCCCGGCGCGTTAGCGGCCCTTCCCCCAGGCCAGGGACGATTATGTATCAATATTGTTGCGTTCGGGCACTCGTGCGAGGGCTCCTGCGGGCTGGGGAGGGGGATCTGGGAATTGGAGGTACGACCGAGATGGCTTGCTCGGGGGGAGGTTTCCTCGCCGAGCAAGCCAGGGTTAGGTGTTGCGCTCTTGACACGTTGTGCATTCTAGGACCCCACTGCTACTCACAACAAGCCCAGTTATGCAGGCCCTGGCCACTCGCCCCTCGGCTATCCGCCCCACCAAGGTGGGCAGGCTCAGCATGTTTGTGCAGCAGAACGATGATGTTTTGTCAGAGTCATGGTTGATCAGCAGATGGCCTCGTTTTGTATTGCTGGCGCCAGTAGTAGGGGCACCGCCGTGGCGTGCAGCTGATGATTGCGCTCAGCGCAGCCGGCGAAGCACAGCGGGCTAGCGAGTCAATGTTTGTCGGCAGCTGCTCCATCGCCAGGCTCTTGGAGCAGCTGCATTCGGAGCTCTTTGTTCTTCACTTGCATTCTGACCAGGGCCGGCCTTGGTGGTGTCCTATCGCAGGCTGCTCGCCGCTCCTCGGTGGTTGTGCGCGCTGATGGCTTCATTGGCTCGTCGACCAACCTGGTGAGCTAGCGTCGTGTGGCGCCGGGAGCCTATTTGCCGAACAATCGTTTCGTTGGAGGCTCCTTCGCGTCCGCAAGTGGCGCTAGCGTGCATGAGCAGCTATGATACAGTCAGATCACACATTCATTCATATGATCTGAGACCTCTCTTGGCTACGGGACCGTGGGTTCCGGGGCACACCGGGTGACCCAACCGGCGCCCGCCTCTTCCCCATGGCTACCCCTCTGGCCACCTAGTCGGAGCTACGCTCCAGCGTGGCGCCCGTGTTTCAGCTGGGAAGCAGTGCATTCAAACAAATGTGACGAGCACTATCGCAAAGGAGCAGCCCAAACCTTGGCCTTAGGAACGCATCCATGACACCTGAATATCGCCTGCCCTTGTGCATGTAGATCATGGTTGCCTCGACCACCGCCACCCTGGCCGCTGCCCGCTTCGGCCTGGCCCCCACCGTGAAGAAGAACACCACCGCTGGCCTGAAGCTGGTGGACTCCAAGAACTCGGCTGGCGTCATCTCCAACGACCCCGCTGGTGAGTCGTTGTCGGCATGGGGTCAGGGCAGGCGGTTGGTGTGGGCGATGCATGTTGGGTGCCGGAGGTGGCTGGGAAAGCTGCAGCGTTACCTGGGGAGGCACTGCAAATAGGTGTCGGGACGGCGCTCTTATCGAGTTATTGGCGGTCTGAGGGGACAAACCCGCCGTAGTGCAGTACAAAAATATGCAAGGACACAGGGCAAGATGTGTACCCAAACGTTAGCTCATTAACGCCGTCACCCTCGGCAATGAAAGCACGAACTAGTCACGTATGTGACCCTTCGCCCGGTCCCCTCTCCGCAGGCTTCACCATTGTGGACGTGCTGGCCATGGGCGCTGCTGGCCACGGTCTGGGCGTTGGCATTGTGCTGGGCCTGAAGGGCATTGGCGCTCTGAACGGAGATCTGGGTGGCTCCGGCGGCCGGATGGTGAGCAAGGGCGAGGAGCTGTTCACCGGCGTGGTGCCCATCCTGGTGGAGCTGGACGGCGACGTGAACGGCCACAAGTTCAGCGTGAGCGGCGAGGGCGAGGGCGACGCCACCTACGGCAAGCTGACCCTGAAGCTGATCTGCACCACCGGCAAGCTGCCCGTGCCCTGGCCCACCCTGGTGACCACCCTGGGCTACGGCCTGCAGTGCTTCGCCCGCTACCCCGACCACATGAAGCAGCACGACTTCTTCAAGAGCGCCATGCCCGAGGGCTACGTGCAGGAGCGCACCATCTTCTTCAAGGACGACGGCAACTACAAGACCCGCGCCGAGGTGAAGTTCGAGGGCGACACCCTGGTGAACCGCATCGAGCTGAAGGGCATCGACTTCAAGGAGGACGGCAACATCCTGGGCCACAAGCTGGAGTACAACTACAACAGCCACAACGTGTACATCACCGCCGACAAGCAGAAGAACGGCATCAAGGCCAACTTCAAGATCCGCCACAACATCGAGGACGGCGGCGTGCAGCTGGCCGACCACTACCAGCAGAACACCCCCATCGGCGACGGCCCCGTGCTGCTGCCCGACAACCACTACCTGAGCTACCAGAGCGCCCTGAGCAAGGACCCCAACGAGAAGCGCGACCACATGGTGCTGCTGGAGTTCGTGACCGCCGCCGGCATCACCCTGGGCATGGACGAGCTGATCAAGGGCGGTGGCGACTACAAGGACCATGACGGTGACTATAAGGATCACGACATCGACTACAAGGACGATGACGACAAGGGCGGCGGCGGCTCGGGCGGCGGCGGCAGCCCCGTGCGCCGCACCGCTATCCCGGCCAACTGGCGTGACGCACTGGGCGGCGGCGGCTCGGGCGGCGGCGGCTCGGGCGGCGGCGGCAGCCCCGTGCGCCGCACCGCTATCCCGGCCAACTGGCGTGACGCACTGGGCGGCGGCGGCTCGGGCGGCGGCGGCTCGGGCGGCGGCGGCAGCCCCGTGCGCCGCACCGCTATCCCGGCCAACTGGCGTGACGCACTGTAAGATCTCTAACTGCACGAATTCTCAGGTCGGGACATCCCCGACCAGGGGCGGCGGGGATGTTGCTGGGCCGATGGAAAGTAGCAACCCAGCCAGCGGCTTCCAGCGCACTCCAGCTGCTCACGGTTGCGACATTGCGCGTGCACGCTTGCGCGTCCCTCACTCGGCCAGCTTGTCGCCGCAGACATCCCTAGCATTGTGCGGACTGCGGTCGTCAGTTAGCGTAGTGGCGGGGCTCAAAGCGTGATGCAGCTGGTGGCTGATTGCATGTGCTACATATGCTGTTATGTTTTGCATGAACTTCGATGCATTGGATGCTGGGTGCACGCGTTTGCATGTGTTTGTGCCGGCATGCTGCCGTCGTCGGCCGTACGTTTACGTTTCTGTGTGCCGGGGTCTTTATTTCCGCCTGCAGACCGACGTCGGTTCCACTAGTTCTAGATGGCAGCAGCTGGACCGCCTGTACCATGGAGAAGAGCTTTACTTGCCGGGATGGCCGATTTCGCTGATTGATACGGGATCGGAGCTCGGAGGCTTTCGCGCTAGGGGCTAGGCGAAGGGCAGTGGTGACCAGGGTCGGTGTGGGGTCGGCCCACGGTCAATTAGCCACAGGAGGATCAGGGGGAGGTAGGCACGTCGACTTGGTTTGCGACCCCGCAGTTTTGGCGGACGTGCTGTTGTAGATGTTAGCGTGTGCGTGAGCCAGTGGCCAACGTGCCACACCCATTGAGAAGACCAACCAACTTACTGGCAATATCTGCCAATGCCATACTGCATGTAATGGCCAGGCCATGTGAGAGTTTGCCGTGCCTGCGCGCGCCCCGGGGGCGAAGCTTGGTACCGCTAGCGTCAGCTGAACTCCTGCTGCCACGCTGACAGAGACGCGCCCATGACGTAGCGCCGACTGCTGCTCCTCTGCTCGTGGAGCTCTGAATCTGCTGCCGGCTGCCCCCGCCCCCGCTGCTGAGGCTGAATCTACTCCGCTCTCCACCCCAGTCCCAGGAAGAGAGGCGCATTTACTTCGCACAGACGTTACAGCACACCCTTGATCATCATCAGCTGCTCTTCCCTGCCGCTGCAACACGCCCCGCGCGATATCCCAGTCGGGAAACCTGTCGTGCCAGCTGCATTAATGAATCGGCCAACGCGCGGGGAGAGGCGGTTTGCGTATTGGGCGCTCTTCCGCTTCCTCGCTCACTGACTCGCTGCGCTCGGTCGTTCGGCTGCGGCGAGCGGTATCAGCTCACTCAAAGGCGGTAATACGGTTATCCACAGAATCAGGGGATAACGCAGGAAAGAACATGTGAGCAAAAGGCCAGCAAAAGGCCAGGAACCGTAAAAAGGCCGCGTTGCTGGCGTTTTTCCATAGGCTCCGCCCCCCTGACGAGCATCACAAAAATCGACGCTCAAGTCAGAGGTGGCGAAACCCGACAGGACTATAAAGATACCAGGCGTTTCCCCCTGGAAGCTCCCTCGTGCGCTCTCCTGTTCCGACCCTGCCGCTTACCGGATACCTGTCCGCCTTTCTCCCTTCGGGAAGCGTGGCGCTTTCTCATAGCTCACGCTGTAGGTATCTCAGTTCGGTGTAGGTCGTTCGCTCCAAGCTGGGCTGTGTGCACGAACCCCCCGTTCAGCCCGACCGCTGCGCCTTATCCGGTAACTATCGTCTTGAGTCCAACCCGGTAAGACACGACTTATCGCCACTGGCAGCAGCCACTGGTAACAGGATTAGCAGAGCGAGGTATGTAGGCGGTGCTACAGAGTTCTTGAAGTGGTGGCCTAACTACGGCTACACTAGAAGAACAGTATTTGGTATCTGCGCTCTGCTGAAGCCAGTTACCTTCGGAAAAAGAGTTGGTAGCTCTTGATCCGGCAAACAAACCACCGCTGGTAGCGGTGGTTTTTTTGTTTGCAAGCAGCAGATTACGCGCAGAAAAAAAGGATCTCAAGAAGATCCTTTGATCTTTTCTACGGGGTCTGACGCTCAGTGGAACGAAAACTCACGTTAAGGGATTTTGGTCATGAGATTATCAAAAAGGATCTTCACCTAGATCCTTTTAAATTAAAAATGAAGTTTTAAATCAATCTAAAGTATATATGAGTAAACTTGGTCTGACAGTTACCAATGCTTAATCAGTGAGGCACCTATCTCAGCGATCTGTCTATTTCGTTCATCCATAGTTGCCTGACTCCCCGTCGTGTAGATAACTACGATACGGGAGGGCTTACCATCTGGCCCCAGTGCTGCAATGATACCGCGAGACCCACGCTCACCGGCTCCAGATTTATCAGCAATAAACCAGCCAGCCGGAAGGGCCGAGCGCAGAAGTGGTCCTGCAACTTTATCCGCCTCCATCCAGTCTATTAATTGTTGCCGGGAAGCTAGAGTAAGTAGTTCGCCAGTTAATAGTTTGCGCAACGTTGTTGCCATTGCTACAGGCATCGTGGTGTCACGCTCGTCGTTTGGTATGGCTTCATTCAGCTCCGGTTCCCAACGATCAAGGCGAGTTACATGATCCCCCATGTTGTGCAAAAAAGCGGTTAGCTCCTTCGGTCCTCCGATCGTTGTCAGAAGTAAGTTGGCCGCAGTGTTATCACTCATGGTTATGGCAGCACTGCATAATTCTCTTACTGTCATGCCATCCGTAAGATGCTTTTCTGTGACTGGTGAGTACTCAACCAAGTCATTCTGAGAATAGTGTATGCGGCGACCGAGTTGCTCTTGCCCGGCGTCAATACGGGATAATACCGCGCCACATAGCAGAACTTTAAAAGTGCTCATCATTGGAAAACGTTCTTCGGGGCGAAAACTCTCAAGGATCTTACCGCTGTTGAGATCCAGTTCGATGTAACCCACTCGTGCACCCAACTGATCTTCAGCATCTTTTACTTTCACCAGCGTTTCTGGGTGAGCAAAAACAGGAAGGCAAAATGCCGCAAAAAAGGGAATAAGGGCGACACGGAAATGTTGAATACTCATACTCTTCCTTTTTCAATATTATTGAAGCATTTATCAGGGTTATTGTCTCATGAGCGGATACATATTTGAATGTATTTAGAAAAATAAACAAATAGGGGTTCCGCGCACATTTCCCCGAAAAGTGCCACCTGACGTCTAAGAAACCATTATTATCATGACATTAACCTATAAAAATAGGCGTATCACGAGGCCCTTTCGTC