Description
The ExcelTaq™ 5X Blood Direct PCR Master Mix Kit is designed for amplifying targeted DNA directly from whole blood, eliminating the need for a lengthy DNA isolation process. ExcelTaq™ Blood Direct PCR Master Mix Kit is a unique blend of 5X concentrated mixture containing all the essential components of a PCR reaction. The ExcelTaq™ Blood Direct PCR Master Mix Kit also contains the PCR-friendly loading and tracking dye solution Orange G. The inclusion of Orange G tracking dye allows for a more convenient post-PCR gel electrophoresis analysis. ExcelTaq™ 5X Blood Direct PCR Master Mix Kit is capable of tolerating the presence of PCR interfering/ inhibiting substances in blood. The ExcelTaq™ 5X Blood Direct PCR Master Mix Kit is ideal for high-throughput screening of blood samples for high reproducibility. ExcelTaq™ 5X Blood Direct DNA PCR Master Mix Kit includes a pair of Positive Control Primers (CCR5) that are compatible with primate blood samples.
Features
5"→3" DNA polymerase activity
No detectable 3"→5" exonuclease (proofreading) activity
Generates PCR products with 3"-dA overhangs
High throughput PCR
High yield PCR
High reproducibility, with reduced pipetting errors
Compatible with most anticoagulants
Applications
Direct amplification of DNA from blood samples
High throughput screening without DNA purification
Suitable for multiplex PCR
Storage
Caution: Avoid Multiple Freeze/Thaw Cycles
4°C for 6 months-20°C for 24 months
Compatible with most anticoagulants
ExcelTaq™ 5X Blood Direct PCR Master Mix amplified 200 bp from differently treated blood using CCR5 specific primers. (M: DM2100)
Contents
Component | Volume |
5X Blood Direct PCR Master Mix | 2 x 1 ml |
Positive Control Primers (10 μM,each) | 25 μl |
CCR5 control primer sequence
CCR5-F:5"-CTCCCAGGAATCATCTTTACC-3"
CCR5-R:5"-TCATTTCGACACCGAAGCAG-3"
Storage
Caution: Avoid Multiple Freeze/Thaw Cycles
4°C for 6 months-20°C for 24 months
Manual
Manual_TP2100_ExcelTaq™ 5X Blood Direct PCR Master Mix Kit
SDS
SDS_TP2100
Recommended PCR Condition
Blood | 3μl |
Forward primer | 0.1– 0.5 µM |
Reverse primer | 0.1– 0.5 µM |
5X Blood Direct PCR Master Mix | 10μl |
H2O | to50 μl |
Total volume | 50µl |
Note: Mix well before PCR Reaction
Recommended PCRProgram
Steps | Temp. | Time | Cycles |
Templatedenature | 94°C | 3 min | 1 |
Denature | 94°C | 30 sec | 25-40 |
Annealing | 50-68°C** | 30 sec | |
Extension | 72°C | 60 sec/kb | |
Final extension | 72°C | 3 min | 1 |
*Optimal PCR condition variesaccording to primers’ thermodynamic properties.
Post amplificationanalysis
Centrifuge PCR reaction mixture at 1000 × g for 1 ~ 3 minutes. Load only the clear supernatant of the PCRproducts for gel electrophoresis analysis.
[TP2000] ExcelTaq™ Blood Direct DNA Polymerase
High yield
Thermostable, up to 50°C, during first strand synthesis
High processivity, generating cDNA up to 8 kb
Reduced RNase H ribonuclease activity
[TP5000] ExcelTaq™ Hot Start II DNA Polymerase
Aptamer-based hot start PCR
Reversible enzyme inactivation
Omits extra enzyme activation step
Convenient for room temperature PCR set-up
High yield and specificity of target amplicons
Wide range of amplicon length (up to 10 kb)
High sensitivity (as low as 1 fg of plasmid)
[TQ1200] ExcelTaq™ 2X Fast Q-PCR Master Mix (SYBR, no ROX)
High Stability
Fast Hot Start
High Sensitivity
Low Background / High Specificity
Suitable for Fast Program
Smart Blue Contrast Dye
[TP1200] ExcelTaq™ 5X PCR Master Dye Mix
5’→3’ DNA polymerase activity
No detectable 3"→5" exonuclease (proofreading) activity
Generates PCR products with 3"-dA overhangs
High yield PCR
High reproducibility
Reduced pipetting errors
Includes tracking dye for direct loading after PCR
ebiomall.com
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求问酶切体系为10ul,内切酶1ul,37℃水浴45min,不知道DNA产物是否完全被切开了?
所用内切酶信息如下
同一管dna有的酶可以切开,比如dra1,ecoR1,Hind111
有的酶又完全不能切动,如BamH1,sac1,xba1,xho1,pst1
请问高手这是什么原因??
配后是否应该分装,因为不能反复冻融?
多谢!
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【】可以这样输入,智能ABC,输入“v1”,后翻3页,就可以看见了,或用紫光拼音中文输入法,输入方括号就是粗体的中括号,实在不行请复制本公告中【】
感谢赐教~!
我做的双酶切反应,酶分别是宝生物的BamHI和XhoI,双酶切体系是:载体及目的片段分别是30ul,通用buffer4ul,XhoI、BamHI各2ul,无核酶水2ul,总体积40ul。载体和目的片段都是经37度酶切6h。载体上面这两个酶切位点的距离是6个碱基,目的片段是由pcr反应获得的,从puc19中p出来的,两端带有这两种酶的酶切位点,酶切完成后,做连接反应,体系如下:目的片段(全长1441bp)4ul,无核酶水3ul,10*T4DNA连接酶缓冲液1ul,载体1ul,T4DNA连接酶1ul,总体积10ul。16度过夜。之后摇菌送沉菌测序结果回来,送了5管一个都不对。重复实验两次还是不对!
之后改为先用BanHI单切载体,体系如下:载体17ul,酶1ul,buffer2ul,总体积20ul,30度切6小时后,Promega纯化试剂盒直接纯化,完了之后再用XhoI,37度酶切6小时,体系如上,再次用promega纯化试剂盒纯化,目的片段用双酶切体系酶切,之后做连接反应,之后铺板,挑取菌落共10个,摇菌13小时,取菌液做pcr鉴定,结果10个菌落全是引物2聚体,跑出来的条带都是100bp左右。
各位老师这是怎么回事呢?为什么连接不上呢?小弟先谢过各位大哥了!