Synthetic Biology Journal ›› 2023, Vol. 4 ›› Issue (1): 30-46.DOI: 10.12211/2096-8280.2022-055
• Invited Review • Previous Articles Next Articles
Yi YANG1, Yufeng MAO1, Chunhe YANG1,2, Meng WANG1, Xiaoping LIAO1, Hongwu MA1
Received:
2022-10-03
Revised:
2022-11-23
Online:
2023-03-07
Published:
2023-02-28
Contact:
Hongwu MA
杨毅1, 毛雨丰1, 杨春贺1,2, 王猛1, 廖小平1, 马红武1
通讯作者:
马红武
作者简介:
杨毅(1986—),男,博士研究生。研究方向为生物信息学、合成生物学、代谢工程。基金资助:
CLC Number:
Yi YANG, Yufeng MAO, Chunhe YANG, Meng WANG, Xiaoping LIAO, Hongwu MA. Recent progress in computational tools for designing editing sequences used in microbial genetic manipulations[J]. Synthetic Biology Journal, 2023, 4(1): 30-46.
杨毅, 毛雨丰, 杨春贺, 王猛, 廖小平, 马红武. 面向微生物遗传操作的编辑序列设计工具的研究进展[J]. 合成生物学, 2023, 4(1): 30-46.
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URL: https://synbioj.cip.com.cn/EN/10.12211/2096-8280.2022-055
Fig. 1 Development of genetic manipulation technologies and corresponding editing sequences and tools for editing sequence design(Blue rectangle: Genetic manipulation technologies; Green rectangle: Editing sequence design requirements; Brown rectangle: Editing sequence design tools; Brown serial numbers indicates that the editing sequence design tools can address the corresponding editing sequence design requirements)
工具 | 基于Primer3 | 特异性检验 | 批量设计 | 功能特色 | 可用性 | URL |
---|---|---|---|---|---|---|
Primer3[ | — | 否 | 否 | 基础引物设计 | 在线/图形界面、离线/命令行 | https://bioinfo.ut.ee/primer3/ |
BatchPrimer3[ | 是 | 否 | 是 | 批量引物设计 | 在线/图形界面 | http://probes.pw.usda.gov/cgi-bin/batchprimer3/batchprimer3.cgi |
ConservedPrimer2.0[ | 是 | 是 | 是 | Sanger测序引物设计 | 在线/图形界面 | https://probes.pw.usda.gov/ConservedPrimers/index.html |
PrimerDesign-M[ | 否 | 是 | 是 | Sanger测序引物设计 | 在线/图形界面 | www.hiv.lanl.gov/content/sequence/PRIMER_DESIGN/primer_design.html |
Primer-BLAST[ | 是 | 是 | 否 | RT-PCR引物设计 | 在线/图形界面 | https://www.ncbi.nlm.nih.gov/tools/primer-blast/ |
PCR designer[ | 否 | 否 | 否 | SNP分析引物设计 | 在线/图形界面 | http://primer1.soton.ac.uk/primer.html |
PRIMEGENS-v2[ | 是 | 否 | 是 | 剪接变体引物设计 | 在线/图形界面、离线/命令行 | http://primegens.org/ |
PrimerSeq[ | 是 | 否 | 是 | 剪接变体引物设计 | 离线/图形界面 | https://primerseq.sourceforge.net/ |
MSP-HTPrimer[ | 是 | 否 | 是 | DNA甲基化分析引物设计 | 离线/命令行 | https://sourceforge.net/projects/msp-htprimer/ |
MPD[ | 否 | 是 | 是 | 多重PCR引物设计 | 离线/命令行 | https://wingolab-org.github.io/mpd-c/ |
Mutation Maker[ | 是 | 是 | 是 | 多重PCR引物设计、氨基酸突变引物设计 | 离线/命令行 | https://github.com/ra100/Mutation_Maker |
PerlPrimer[ | 否 | 否 | 是 | Sanger测序引物设计、RT-PCR引物设计、开放阅读框(ORF)搜索及相关引物设计 | 离线/图形界面 | https://perlprimer.sourceforge.net/ |
Table 1 Major primer design tools with different functions
工具 | 基于Primer3 | 特异性检验 | 批量设计 | 功能特色 | 可用性 | URL |
---|---|---|---|---|---|---|
Primer3[ | — | 否 | 否 | 基础引物设计 | 在线/图形界面、离线/命令行 | https://bioinfo.ut.ee/primer3/ |
BatchPrimer3[ | 是 | 否 | 是 | 批量引物设计 | 在线/图形界面 | http://probes.pw.usda.gov/cgi-bin/batchprimer3/batchprimer3.cgi |
ConservedPrimer2.0[ | 是 | 是 | 是 | Sanger测序引物设计 | 在线/图形界面 | https://probes.pw.usda.gov/ConservedPrimers/index.html |
PrimerDesign-M[ | 否 | 是 | 是 | Sanger测序引物设计 | 在线/图形界面 | www.hiv.lanl.gov/content/sequence/PRIMER_DESIGN/primer_design.html |
Primer-BLAST[ | 是 | 是 | 否 | RT-PCR引物设计 | 在线/图形界面 | https://www.ncbi.nlm.nih.gov/tools/primer-blast/ |
PCR designer[ | 否 | 否 | 否 | SNP分析引物设计 | 在线/图形界面 | http://primer1.soton.ac.uk/primer.html |
PRIMEGENS-v2[ | 是 | 否 | 是 | 剪接变体引物设计 | 在线/图形界面、离线/命令行 | http://primegens.org/ |
PrimerSeq[ | 是 | 否 | 是 | 剪接变体引物设计 | 离线/图形界面 | https://primerseq.sourceforge.net/ |
MSP-HTPrimer[ | 是 | 否 | 是 | DNA甲基化分析引物设计 | 离线/命令行 | https://sourceforge.net/projects/msp-htprimer/ |
MPD[ | 否 | 是 | 是 | 多重PCR引物设计 | 离线/命令行 | https://wingolab-org.github.io/mpd-c/ |
Mutation Maker[ | 是 | 是 | 是 | 多重PCR引物设计、氨基酸突变引物设计 | 离线/命令行 | https://github.com/ra100/Mutation_Maker |
PerlPrimer[ | 否 | 否 | 是 | Sanger测序引物设计、RT-PCR引物设计、开放阅读框(ORF)搜索及相关引物设计 | 离线/图形界面 | https://perlprimer.sourceforge.net/ |
工具 | 基于Primer3 | 支持的DNA片段组装技术 | 批量设计 | 可用性 | URL |
---|---|---|---|---|---|
J5[ | 是 | Golden Gate/Gibson/ CPEC/SLIC | 是 | 在线/图形界面 | https://j5.jbei.org/ |
Geneious | 否 | Gibson | 是 | 在线/图形界面 | https://www.geneious.com/ |
Raven[ | 否 | Gibson | 是 | 在线/图形界面 | http://ravencad.org/ |
MCDS[ | 否 | GoldenGate/Gibson/ SLIC/Gateway | 否 | 离线/命令行 | https://github.com/errisy/MCDS |
NEBbridge Golden Gate[ | 未知 | Golden Gate | 是 | 在线/图形界面 | https://goldengate.neb.com/ |
iBioCAD GGA[ | 未知 | Golden Gate | 是 | 在线/图形界面 | https://ibiocad.igb.illinois.edu/ |
PlasmidMaker[ | 是 | Golden Gate/Gibson | 是 | 在线/图形界面 | https://biofoundry.web.illinois.edu/ |
Table 2 Representative design tools for DNA assembly
工具 | 基于Primer3 | 支持的DNA片段组装技术 | 批量设计 | 可用性 | URL |
---|---|---|---|---|---|
J5[ | 是 | Golden Gate/Gibson/ CPEC/SLIC | 是 | 在线/图形界面 | https://j5.jbei.org/ |
Geneious | 否 | Gibson | 是 | 在线/图形界面 | https://www.geneious.com/ |
Raven[ | 否 | Gibson | 是 | 在线/图形界面 | http://ravencad.org/ |
MCDS[ | 否 | GoldenGate/Gibson/ SLIC/Gateway | 否 | 离线/命令行 | https://github.com/errisy/MCDS |
NEBbridge Golden Gate[ | 未知 | Golden Gate | 是 | 在线/图形界面 | https://goldengate.neb.com/ |
iBioCAD GGA[ | 未知 | Golden Gate | 是 | 在线/图形界面 | https://ibiocad.igb.illinois.edu/ |
PlasmidMaker[ | 是 | Golden Gate/Gibson | 是 | 在线/图形界面 | https://biofoundry.web.illinois.edu/ |
工具 | 编辑效率预测方法 | 特异性预测方法 | PAM | 支持的改造类型 | URL |
---|---|---|---|---|---|
E-CRISP[ | 假设先验 | 假设先验 | NGG | 敲除、激活、抑制 | http://www.e-crisp.org/E-CRISP/index.html |
CRISPR-ERA[ | 假设先验 | 假设先验 | NGG | 敲除、激活、抑制 | http://CRISPR-ERA.stanford.edu |
EuPaGDT[ | 假设先验 | 假设先验 | NGG、TTTN、NGA等 | 敲除 | http://grna.ctegd.uga.edu/ |
CRISPOR[ | 假设先验/机器学习 | 假设先验 | NGG、TTTN、NGA等 | 敲除 | http://crispor.tefor.net/ |
CHOPCHOP[ | 假设先验/机器学习 | 假设先验 | NGG、TTTN、NGA等 | 敲除、敲入、激活、抑制 | http://chopchop.cbu.uib.no/ |
CRISPETa[ | 假设先验 | 数据库搜索 | NGG | 敲除 | http://crispeta.crg.eu |
CRISPR-P[ | 假设先验 | 假设先验 | NGG、NRG、TTN等 | 敲除 | http://crispr.hzau.edu.cn/CRISPR2/ |
CRISPRscan[ | 机器学习 | 假设先验 | NGG、TTTN、NGN等 | 敲除 | https://www.crisprscan.org |
WU-CRISPR[ | 机器学习 | 假设先验 | NGG | 敲除 | http://crisprdb.org/wu-crispr/ |
CRISTA[ | 机器学习 | 机器学习 | NGG | 敲除 | https://crista.tau.ac.il/ |
Elevation[ | 机器学习 | 机器学习 | NGG | 敲除 | https://crispr.ml/ |
Table 3 Representative hypothesis-driven and learning-based sgRNA design tools
工具 | 编辑效率预测方法 | 特异性预测方法 | PAM | 支持的改造类型 | URL |
---|---|---|---|---|---|
E-CRISP[ | 假设先验 | 假设先验 | NGG | 敲除、激活、抑制 | http://www.e-crisp.org/E-CRISP/index.html |
CRISPR-ERA[ | 假设先验 | 假设先验 | NGG | 敲除、激活、抑制 | http://CRISPR-ERA.stanford.edu |
EuPaGDT[ | 假设先验 | 假设先验 | NGG、TTTN、NGA等 | 敲除 | http://grna.ctegd.uga.edu/ |
CRISPOR[ | 假设先验/机器学习 | 假设先验 | NGG、TTTN、NGA等 | 敲除 | http://crispor.tefor.net/ |
CHOPCHOP[ | 假设先验/机器学习 | 假设先验 | NGG、TTTN、NGA等 | 敲除、敲入、激活、抑制 | http://chopchop.cbu.uib.no/ |
CRISPETa[ | 假设先验 | 数据库搜索 | NGG | 敲除 | http://crispeta.crg.eu |
CRISPR-P[ | 假设先验 | 假设先验 | NGG、NRG、TTN等 | 敲除 | http://crispr.hzau.edu.cn/CRISPR2/ |
CRISPRscan[ | 机器学习 | 假设先验 | NGG、TTTN、NGN等 | 敲除 | https://www.crisprscan.org |
WU-CRISPR[ | 机器学习 | 假设先验 | NGG | 敲除 | http://crisprdb.org/wu-crispr/ |
CRISTA[ | 机器学习 | 机器学习 | NGG | 敲除 | https://crista.tau.ac.il/ |
Elevation[ | 机器学习 | 机器学习 | NGG | 敲除 | https://crispr.ml/ |
Fig. 4 Schematic diagram for CRISPR/Cas-based homologous recombination and its editing sequencesDF—Deleted fragment; P — Primer; UHAmin—Minimum length of upstream homology arm; UHAmax— Maximum length of upstream homology arm; ODRUHA—Optional design region of the upstream primer of the upstream homologous arm; DHAmin —Minimum length of downstream homology arm; DHAmax— Maximum length of downstream homology arm; ODRDHA— Optional design region of the downstream primer of downstream homologous arm; R—sgRNA (dark green and blue of the sgRNA border lines represent on-target efficiency and off-target risk, respectively); TP— Test primer; UHA— Upstream homologous arm; DHA— Downstream homologous arm; USSmin—Minimum length of upstream spacer sequence (sequence from the 3'-end of the upstream verification primer to the 5'-end of the upstream homology arm); USSmax— Maxmum length of upstream spacer sequence; ODRUSS— Optional design region of the upstream verification primer; DSSmin— Minimum length of downstream spacer sequence (sequence from the 3'-end of the downstream verification primer to the 3'-end of the downstream homology arm); DSSmax— Maxmum length of downstream spacer sequence; ODRDSS—Optional design region of the downstream verification primer
工具 | 编辑序列设计 | 遗传改造类型 | 是否高通量 | 可支持物种 | 可编辑位点 | web应用 |
---|---|---|---|---|---|---|
Yeastriction[ | 引物设计sgRNA设计 | 删除 | 否 | Saccharomyces cerevisiae(33个菌株) | 已被注释的基因 | http://yeastriction.tnw.tudelft.nl/#!/ |
GeneTargeter[ | 引物设计sgRNA设计 | 删除、表达弱化 | 是 | Plasmodium falciparum | 已被注释的基因 | http://genetargeter.mit.edu/ |
GEDpm-cg[ | 引物设计、同源臂设计 | 单点突变 | 是 | Corynebacterium glutamicum | 基因组任意区域 | https://gedpm-cg.biodesign.ac.cn/ |
AutoESD[ | 引物设计、同源臂设计 | 插入、删除、替换、单点突变 | 是 | 多种原核、真核微生物 | 基因组任意区域 | https://autoesd.biodesign.ac.cn/ |
Table 4 Editing sequence design tools for the whole workflow of genome editing
工具 | 编辑序列设计 | 遗传改造类型 | 是否高通量 | 可支持物种 | 可编辑位点 | web应用 |
---|---|---|---|---|---|---|
Yeastriction[ | 引物设计sgRNA设计 | 删除 | 否 | Saccharomyces cerevisiae(33个菌株) | 已被注释的基因 | http://yeastriction.tnw.tudelft.nl/#!/ |
GeneTargeter[ | 引物设计sgRNA设计 | 删除、表达弱化 | 是 | Plasmodium falciparum | 已被注释的基因 | http://genetargeter.mit.edu/ |
GEDpm-cg[ | 引物设计、同源臂设计 | 单点突变 | 是 | Corynebacterium glutamicum | 基因组任意区域 | https://gedpm-cg.biodesign.ac.cn/ |
AutoESD[ | 引物设计、同源臂设计 | 插入、删除、替换、单点突变 | 是 | 多种原核、真核微生物 | 基因组任意区域 | https://autoesd.biodesign.ac.cn/ |
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