A common bottleneck for genome editing applications involves a lack of efficient methods for detecting successfully engineered SNPs or insertions. While many researchers rely on sequencing to verify editing, this typically requires isolation and expansion of single-cell clones and yields data that can be challenging to analyze. The Guide-it Knockin Screening Kit provides a streamlined method for identifying precise edits at any locus, either in heterogeneous or clonal cell populations. The assay takes only 4 hours to complete and can be performed using basic laboratory equipment.
A common bottleneck for genome editing applications involves a lack of efficient methods for detecting successfully engineered SNPs or insertions. While many researchers rely on sequencing to verify editing, this typically requires isolation and expansion of single-cell clones and yields data that can be challenging to analyze. The Guide-it Knockin Screening Kit provides a streamlined method for identifying precise edits at any locus, either in heterogeneous or clonal cell populations. The assay takes only 4 hours to complete and can be performed using basic laboratory equipment.
One of the most powerful applications of genome editing technologies such as the CRISPR/Cas system is the ability to introduce precise changes at genomic loci of interest. However, the likelihood of success for this type of application is generally low because it often relies on an endogenous repair mechanism known as homology-directed repair (HDR) that occurs at relatively low frequency. Consequently, there are two different stages when the detection of successful HDR events is critically important. The first stage involves optimization of experimental conditions to achieve the highest percentage of error-free HDR events in an edited population before moving forward with the isolation of single-cell clones. The second stage involves identification of cell lines carrying the edit of interest after single-cell isolation and expansion in 96-well plates. The Guide-it Knockin Screening Kit addresses the need for sensitive detection of successful HDR at both stages by providing a simple fluorescence-based method that can be applied to screen edited populations as well as clones from 96-well plates for edits ranging in size from single-nucleotide substitutions to longer insertions.
The kit assay consists of PCR amplification of the genomic target site, followed by an enzymatic assay employing assay-specific oligos that can be designed with our online tool. The dual-color fluorescence-based readout from the assay can be measured using a standard plate reader or qPCR machine. A positive fluorescent signal from the assay is highly correlated with the correct introduction of the desired edit. For engineering SNPs, the assay enables detection of single-nucleotide substitutions with high sensitivity in both mixed and clonal populations and can be used to positively identify heterozygous clones carrying one copy each of the edited (SNP) and unedited (WT) alleles. For scenarios involving knockin of longer sequences, the assay allows for the simultaneous detection of seamless insertions at both 5" and 3" ends of the incorporated sequence.
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试剂盒在全球市场上的研发与销售呈快速上升趋势,2005年全球市场销售额超过200亿美元,且以15%左右的速度逐年增长.一方面是试剂盒的迅猛发展,而另一方面试剂盒市场良莠不齐的现象愈加明显,试剂盒的生产、销售及认证认可体制尚不完善,没有相应的标准或质量评价政策.且其灵敏度,稳定性及假阴/阳性控制尚不能满足检测需要,采用试剂盒进行检测的公信度受到质疑.
同时,食品安全领域是当前问题最多、最受关注的领域,这个领域的检测包括了物理、化学、微生物及分子生物学基础理论,无论是按检测原理、用途还是其它分类方式,涉及食品安全检测项目的试剂盒的品种是最全最多的.因此,从该领域着手从事评价制度的研究,便于获得基础性数据结果,并由此推广至动植物检疫及其它领域.
随着H7N9禽流感疫情的不断发散,国家流感中心已经多次发放人感染H7N9禽流感检测试剂,覆盖了全国31个流感网络实验室,并表示,诊断试剂的广泛发放是实现关口前移,控制疫情传播、蔓延的重要手段.而一旦H7N9监测关口继续前移,主动监测范围扩大,病毒检测试剂的需求量将进一步加大.
可采用H7N9亚型禽流感病毒RNA检测试剂盒（荧光PCR法）和H7N9禽流感病毒核酸检测试剂盒（PCR-荧光探针法）,定价分别为48人份/盒和48反应/盒,相比市场此前预期的100-200元之间的价格定位低了很多.在检验方法上,卫纪委提醒,前者需要配备全自动荧光PCR检测仪专用PCR扩增管和核酸分离试剂盒(硅胶膜吸附法)等必须设备及咽拭子样本,后者卫计委推荐采用达安基因生产的核酸提取试剂盒进行检验.
ldh试剂盒指乳酸脱氢酶检测试剂盒，是一种基于diaphorase催化的INT显色反应，通过比色法检测细胞毒性时释放的乳酸脱氢酶活性或检测其它样品中的乳酸脱氢酶活性的试剂盒。可以用于常规的乳酸脱氢酶活性的检测，更常用于以LDH释放为指标的细胞毒性检测。
ctl毒性杀伤检测试剂盒是基于LDH在胞浆内含量丰富，正常时不能通过细胞膜，当细胞受损伤或死亡时可释放到细胞外，此时细胞培养液中LDH活性与细胞死亡数目成正比，用比色法测定并与靶细胞对照孔LDH活性比较，可计算效应细胞对靶细胞的杀伤。
我用Promega公司的双荧光素酶检测试剂盒（E2920）检测到的firefly萤光素酶活性很低，只有4*10的3次方；海肾萤光素酶活性有10的6次方。
我用Ad293细胞做了转染，fireflyluciferase质粒：RanillaLuciferase质粒=0.1ug：0.025ug/一个孔（96孔板），共转染了二天，再进行双萤光素酶检测。
我想了解fireflyluciferase活性用promega的这个E2920-双萤光素酶试剂盒检测得到10的3次方，这种数值正常吗？
我做了3次重复实验，每次firefly萤光素酶活性很低，只有4*10的3次方；而海肾萤光素酶活性有10的6次方。
求指教？fireflyluciferase活性低？会是什么原因呢？
实验室要开展支原体检测，方法是PCR法，先要采购试剂盒，用过的同学给推荐一下好用的品牌呗
