{"product_id":"m-mlv-reverse-transcriptase","title":"M-MLV Reverse Transcriptase","description":"\u003cp\u003e\u003cstrong style=\"font-size: 0.875rem;\"\u003eThe MMLV \u003c\/strong\u003e\u003cspan style=\"font-size: 0.875rem;\"\u003eReverse Transcriptase(short name MMLV) i is iolated from E.coli carrying rat leukemia virus pol gene consists of a single peptide, molecule weight 71kd.It possesses RNA and DNA depended polymerase activity and weak RNase H activity.\u003c\/span\u003e\u003cbr\u003e\u003c\/p\u003e\n\u003cp\u003e\u003cstrong\u003eConcentration: \u003c\/strong\u003e 200U\/μl\u003cbr\u003e\u003cstrong\u003eComponent : \u003c\/strong\u003e M-MLV （200U\/μl） 5×Buffer （with DTT）\u003cbr\u003e\u003cstrong\u003ePackage：\u003c\/strong\u003e Bulk\u003c\/p\u003e\n\u003cp\u003e\u003cstrong\u003eFeatures:\u003c\/strong\u003e\u003cbr\u003eWeak RNaseH activity; High cDNA yield\u003cbr\u003e\u003cstrong\u003eStoreage: \u003c\/strong\u003e-20℃\u003cbr\u003eApplication：Synthesis of the first chain cDNA, cDNA Library construction, one-step RT-PCR, primer extention, 3′ and 5′RACE\u003cbr\u003e\u003cstrong\u003eSource:\u003c\/strong\u003e Recombination of E.coli containing Moloney murine leukemia virus reverse transcriptase gene from clone of Moloney murine .\u003cbr\u003e\u003cstrong\u003eUnit definition:\u003c\/strong\u003eOne unit is defined as the required enzyme incorporate 1 nm dNTP into a polynucleotide fraction in 10 min at 37℃, taking polyA﹒poly(dT)12-18 as template-primer.\u003cbr\u003e\u003cstrong\u003eProcedure:\u003c\/strong\u003e\u003cbr\u003e1. add the next reaction mixture to ice bath tube :\u003cbr\u003e1) template RNA\u003cbr\u003etotal RNA 0.1-5μg\u003cbr\u003eor total poly(A)+mRNA 0.1-0.5μg\u003cbr\u003eor unique RNA 0.01pg-0.5μg\u003cbr\u003e2) primer\u003cbr\u003eOligo(dT)18 (0.5μg\/μl) 1μl\u003cbr\u003eOr stochastic primer（0.2μg\/μl） 1μl\u003cbr\u003eOr sequence especially primer 20pmol\u003cbr\u003e3) RNase-free ddh2o : constant volume to 11μl\u003cbr\u003e2. Gently mix and water bath for 5 min in 70℃ and chill on ice.\u003cbr\u003e3. Put the tube into ice and add the next composition :\u003cbr\u003e5×Reaction Buffer 4μl\u003cbr\u003eRNase Inhibitor (40U\/μl) 0.5μl\u003cbr\u003edNTP Mix(10mmol\/L) 2μl\u003cbr\u003eadd water to 19ul ,gently mix and then water bath for 5 min in 37℃ ; or for 5 min in 25℃ for random primer \u003cbr\u003e4. Spin down for a few seconds. Add 1μl M-MLV RT（200U\/μl）\u003cbr\u003e5. Incubate at 42℃ for 60min(if use a random primer,first incubate for 10min in 25℃\u003cbr\u003e6. Inactivate at 70℃ for 10min.\u003cbr\u003e\u003cstrong\u003ePCR Reaction\u003c\/strong\u003e\u003cbr\u003e1. Transfer 10% volume of first reaction solution (2 μl ) to a proper PCR tube .\u003cbr\u003enote: the first reaction solution can be directly used as PCR template without purification ,the dosage is about 1-5μl. if excessively used ,the salt and Random primers in first reaction solution will restrain the activity of DNA polymerase .if purification needed ,it can follow the next :after reaction end of cDNA synthesis (step 6) , add RNase A in reaction system , 10 min in 37℃ ,use DP1501 recover cDNA .\u003cbr\u003e2. add next solution by order .\u003cbr\u003e5μl 10X PCR Buffer\u003cbr\u003e1μl 10mM dNTP mix\u003cbr\u003e1μl 10μM Primer #1 (customer supplied)\u003cbr\u003e1μl 10μM Primer #2 (p customer supplied)\u003cbr\u003exμl H20 (total reaction volume:49μl)\u003cbr\u003e1μl Taq DNA polymerase\u003cbr\u003e3. Mix thoroughly and add 50μl mineral oil to the surface of liquid.\u003cbr\u003e4. Amplified reaction : according to annealing temperature or gene copy number or technical parameter of Taq DNA polymerase , setting amplified condition , specify reference to specification of DNA polymerase ,the usually cycle number is 30-35\u003cbr\u003e5 Detect the product in agarose containing EB or another fluorescent dye.\u003c\/p\u003e","brand":"Gene Bio Systems","offers":[{"title":"5000U","offer_id":39766547136612,"sku":"R1041","price":11200.0,"currency_code":"JPY","in_stock":true},{"title":"25000U","offer_id":39903278661732,"sku":"R1042","price":48700.0,"currency_code":"JPY","in_stock":true},{"title":"50000U","offer_id":42761391505508,"sku":"R1043","price":89000.0,"currency_code":"JPY","in_stock":true}],"thumbnail_url":"\/\/cdn.shopify.com\/s\/files\/1\/0558\/8588\/9636\/files\/SealingFilmGuide.png?v=1765475529","url":"https:\/\/www.genebiosystems.com\/en-jp\/products\/m-mlv-reverse-transcriptase","provider":"GeneBio ","version":"1.0","type":"link"}