Simple and Effective Squash-PCR for Rapid Genotyping of Industrial Microalgae
Abstract
:1. Introduction
2. Materials and Methods
2.1. Strains
2.2. Squash Preparation
2.3. Genomic DNA Extraction
2.4. PCR Procedure
2.5. Primers for PCR
2.6. Microscopy
2.7. Cell Counting
3. Results
3.1. DNA Template Preparation via Squash Technique
3.2. Efficacy of Squash-PCR in Diverse Microalgae Strains
3.3. Evaluation of Squash-PCR in Microalgae Using Different Cell Numbers and Primer Pairs
4. Discussion
5. Conclusions
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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No. | Microalgae Species | 1 Direct-PCR | Squash-PCR |
---|---|---|---|
1 | Monoraphidium minutum 26B-AM | + | ++ |
2 | Chlorella sorokiniana DOE1412 | + * | ++ |
3 | Tetraselmis striata LANL1001 | − | ++ |
4 | Picochlorum celeri TG2 | + * | ++ |
5 | Scenedesmus obliquus UTEX 393 | + | ++ |
6 | Chlamydomonas reinhardtii UTEX 89 | + | ++ |
7 | Scenedesmus sp. UTEX 1589 | + * | ++ |
8 | Haematococcus pluvialis UTEX 2505 | + | ++ |
9 | Chlorella vulgaris UTEX 395 | + | ++ |
10 | Chlamydomonas reinhardtii UTEX 90 | + | ++ |
11 | Botryococcus braunii UTEX 572 | + | ++ |
12 | Dunaliella salina UTEX LB 200 | − | ++ |
Primer | Sequence | Microalgae Species | Gene | Size (bp) |
---|---|---|---|---|
102_F | AGGAGAAGTCGTAACAAGGT | Strains 1 to 12 | ITS2 | 700–900 |
103_R | TCCTCCGCTTATTGATATGC | |||
154_F | TTGCACACAAGAACGCATGA | Chlamydomonas reinhardtii | FTSY | 815 |
155_R | GGCATTGGAGTAAACGACCC | |||
156_F | CTACGTCGCCTTACTGTGTG | Chlamydomonas reinhardtii | ZEP | 809 |
157_R | GATTGCCTACTCACCACTCG | |||
197_F | ATGGTTCCACAAACTGAAACG | Dunaliella salina | rbcL | 480 |
198_R | GTCACGCTCAACTTGAATACC | |||
199_F | CGTAGACTGTGTAGAAGCTG | Dunaliella salina | orf121 | 518 |
200_R | GTGGAACTACACCTTCAGGT | |||
201_F | ATGCCAATTGGTGTTCCAC | Dunaliella salina | clpP | 496 |
202_R | GTGCATTAAAATCGTAAGGCG | |||
203_F | CAATGCGCACACCAGAAG | Dunaliella salina | atpA | 551 |
204_R | GCATATAGTTTCCTGCGTCC | |||
205_F | ATGGCACGTGCTAAATTTGA | Dunaliella salina | tufA | 524 |
206_R | GCAGAACCTGAAACGATAGG |
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Yuan, G.; Gao, S.; Czajka, J.J.; Dai, Z.; Pomraning, K.R.; Duong, R.D.; Hofstad, B.A.; Deng, S. Simple and Effective Squash-PCR for Rapid Genotyping of Industrial Microalgae. Life 2024, 14, 115. https://doi.org/10.3390/life14010115
Yuan G, Gao S, Czajka JJ, Dai Z, Pomraning KR, Duong RD, Hofstad BA, Deng S. Simple and Effective Squash-PCR for Rapid Genotyping of Industrial Microalgae. Life. 2024; 14(1):115. https://doi.org/10.3390/life14010115
Chicago/Turabian StyleYuan, Guoliang, Song Gao, Jeffrey J. Czajka, Ziyu Dai, Kyle R. Pomraning, Rylan D. Duong, Beth A. Hofstad, and Shuang Deng. 2024. "Simple and Effective Squash-PCR for Rapid Genotyping of Industrial Microalgae" Life 14, no. 1: 115. https://doi.org/10.3390/life14010115